Background: Child malnutrition is a major epidemiological problem in developing countries, especially in African countries. Nutrition education for mothers can alleviate this malnutrition in their young children. The objective of this study was to make a systematic review to assess the effect of intervention programs in nutrition education for African mothers on the nutritional status of their infants. Methods: A bibliographic search was carried out in the PubMed database for clinical trials between November 2012 and 2021. The studies should contain educational programs to evaluate the impact on the infant’s nutritional indicators in children under 5 years . Results: A total of 20 articles were selected, of which 53% evaluated infant’s food consumption, 82% anthropometric measurements and 30% nutritional knowledge. In general, nutritional education programs are accredited with some significant improvements in food and nutrient consumption, knowledge and dietary practices in complementary feeding, but only those studies that implemented strategies in agriculture, educational workshops and supplementation obtained reductions in chronic malnutrition figures. Limitations: There is high heterogeneity in the articles included, since the intervention programs have different approaches. Conclusions: Programs that implemented actions of national agriculture or nutritional supplementation reap the greatest benefits in curbing infant malnutrition. Child malnutrition is a worrying public health problem in developing countries. According to 2019 statistics, it is estimated that worldwide, one in five children under 5 years of age has some degree of malnutrition, about 150 million suffer from insufficient stature and almost 50 million are wasted . ChildreOn the other hand, it has been observed that good health care practices, better nutrition and living conditions and greater general knowledge of the mother have a positive effect on maternal and child health , while tVarious studies have suggested that providing mothers with educational strategies about feeding has a significant positive impact on their children’s nutritional status, especially when they are integrated into the local contexts they are designed for. Dewey and Adu-Afarwuah in a systematic review (2008)  evaluateThe present study aims to carry out a systematic review to assess the effect of intervention programs in nutrition education, designed for African mothers, on the nutritional status of children of infant age.Following PRISM Systematic Review Guidelines, a bibliographic search was carried out in the PubMed database of clinical trials published between November 2012 and 2021, the last date of search being 15 June 2021, using the MESH terms with the following combination:(((programs OR intervention) AND education AND ) AND  AND (child*))) AND (Africa OR “Africa South of the Sahara”[Mesh] OR “South Africa”[Mesh] OR “Mozambique”[Mesh]).Clinical trials were included that carried out intervention programs in nutrition education for mothers or caregivers of children less than 5 years of age who were African and malnourished. Those focused on educational programs were to be selected, as well as those that, in addition to being educational, are complemented with the promotion of domestic agriculture, food/nutrient supplementation or similar characteristics.Moreover, the programs had to be evaluated by indicators that assessed the knowledge and skills acquired by mothers or caregivers about infant feeding, infants’ anthropometric development and nutritional status .We did not apply any language restriction.Manuscripts that did not have the above characteristics were excluded, as were those focused on children or mothers with specific diseases , non-health education-based programs, non-Africans and children over 5 years of age.All data from the eligible clinical trials were independently abstracted by two researchers (BC and VA) without significant disagreements. The screening of titles and abstracts for all studies was independently performed by two authors (BC and VA) in order to identify potentially relevant articles. Likewise, these two authors (BC and VA) performed full-text screenings and with mutual consensus, confirmed that studies met the study’s inclusion and exclusion criteria.Subsequently, the bibliographic citations referenced in the articles selected were reviewed to locate other investigations (CJ). The authors resolved discrepancies after discussion articles were not excluded for reasons of language.Nutrition & Dietetics.The quality of the studies selected was examined by the ranking of the journals published through the web InCites Journal Citation Reports and referenced by quartiles (Q) in In data management, each study was taken and evaluated by the year of publication and the country where it was carried out, the educational guides used and type of intervention , sample size and duration of the intervention, evaluated indicators and quantification of the results. Every study should evaluate baseline vs. final line; we took the results with a significance value lower than 0.05.A total of 122 articles were retrieved from the PudMed search. From these, 41 were selected on the strength of the title, of which 10 were excluded due to the content of the abstract and 11 after a complete reading of the text. Subsequently, five studies were included after a manual search to complete a selection of 20 articles see .Virtually all of the selected studies are randomized clinical trials, except for three ,45,52, wSome of the nutrition education programs were implemented by mothers who are leaders in the environment ,45 and oThe effect of the program on mothers or infants was assessed by various indicators. Several EPs have used different questionnaires to evaluate eating activities and the knowledge acquired by the mothers in food hygiene, food selection and complementary feeding ,24,45,47Although there are a considerable number of nutritional interventions in Africa to alleviate infant malnutrition, this review has found few clinical trial-based nutrition education intervention programs. Although our analysis has shown some beneficial effect in educational programs on the nutritional status of children, it is not enough to generate nutritional changes in children. In order to observe which programs have had better results, the discussion was structured on the effect observed on the analysed result variables, first, over knowledge and feeding practices acquired by mothers, together with the nutritional intake carried out by the infant, and second, on the infant’s anthropometric development and degree of malnutrition.The main objective of advanced studies on nutrition education is to bring about a change in the attitudes and beliefs of African mothers about infant feeding and to increase their understanding ,14,46. DIn this review, we try to analyse the feeding practices and nutritional knowledge of the different types of programs. We found that the food and nutritional intake of infants in the EP was evaluated by analysing nutritional intake and using the MMF and MDD. However, only two clinical trials conducted in Ethiopia and Kenya analysed nutritional intake ,45. In tp ≤ 0.001), proteins  and iron  in children from the intervention groups  also indicates the need for coordinated policy initiatives across the entire food system, from food production to marketing and consumer demand, to identify more effective systems .The Foresight Africa project consists of a series of reports on the main points to take into account in the development of effective policies to reduce malnutrition on this continent, explaining the points they consider to be priorities. These reports highlight the importance of coordinating scientific evidence with the activities of farmers to generate more effective solutions, adapted to the demand of the community. They also emphasize the emphasis on the application of digital technologies in food production, which allow preventing climate crises, improving irrigation systems, etc., as well as improving the use and safety of food, among others. These reports also consider that it is not possible to address food insecurity without addressing the causes of conflict and fragility related to agriculture and recommend strengthening the capacity of local institutions to promote the design and implementation of community-based approaches, making better use of natural resources in these fragile areas .Although the studies on nutrition education to combat child malnutrition in Africa found common ground, joint evaluation of the results of the different programs is difficult given their methodological diversity and outcome variables. However, it should be noted that although the interventions on nutrition education carried out on mothers and caregivers were useful for improving dietary diversity and attitudes toward recommended feeding, they appear to be insufficient to generate nutritional changes in children. For this reason, the programs should be accompanied by actions of national agriculture or nutritional supplementation. Only then will people be able to adhere to the recommendations and guidelines of a healthy diet to stop the processes of child malnutrition since food shortages are a latent problem in Africa. Despite the observed benefits, more studies are required to provide more evidence on the characteristics of the most effective interventions.


This review contains a personal account of the role played by the PDB in the development of the field of molecular chaperones and protein homeostasis, from the viewpoint of someone who experienced the concurrent advances in the structural biology, electron microscopy, and chaperone fields. The emphasis is on some key structures, including those of Hsp70, GroEL, Hsp90, and small heat shock proteins, that were determined as the molecular chaperone concept and systems for protein quality control were emerging. These structures were pivotal in demonstrating how seemingly nonspecific chaperones could assist the specific folding pathways of a variety of substrates. Moreover, they have provided mechanistic insights into the ATPase machinery of complexes such as GroEL/GroES that promote unfolding and folding and the disaggregases that extract polypeptides from large aggregates and disassemble amyloid fibers. The PDB has provided a framework for the current success in curating, evaluating, and distributing structural biology data, through both the PDB and the EMDB. The elucidation of the heat shock response in the 1960s, showing that a set of characteristic changes in gene expression is triggered by environmental changes such as heat stress . The structure was published in 1990, and the PDB entry (3hsc) appeared in 1995 . But it would take until 2012 for the two domains to be captured together, tightly interacting in a dramatically changed conformation, instead of two separate structures loosely tethered by a flexible linker . Opening and shutting of the substrate-trapping lid is allosterically coupled to the ATPase cycle  to look at large complexes. We were still in the era when disdain was a typical reaction of crystallographers to “blobology,” if they had indeed even heard of using EM to look at macromolecular complexes. Soon after I set foot in the department, Steve told me about the large protein he was trying to crystallize that was somehow involved in protein assembly, and I was intrigued by the available EM images. I was easily persuaded to have a look, and from my first glimpse of it I was totally captivated. I felt sure that we could learn a lot from the images. The pioneering work on bacteriorhodopsin by Henderson and Unwin . The structure showed that the internal cavities were lined by hydrophobic sites, and targeted mutagenesis showed the key role of the hydrophobic sites in substrate binding and folding . The key elements of the mechanism were becoming clear: the nonnative protein was trapped on the hydrophobic lining of an open cavity, but then the combined actions of ATP and GroES binding caused a major reorganization of the complex so that the substrate was ejected from its binding sites but then trapped inside an enclosed cavity, capped by GroES, with a now hydrophilic lining—the folding chamber . This encapsulation, possibly following some forced unfolding during the dramatic restructuring of the complex . Subsequent ATP hydrolysis allowed the release of the GroES lid and the contents of the folding chamber, whether folded or not. Nonnative protein would be recaptured for another round of interaction, whereas native protein would not incorrectly expose hydrophobic surface and would no longer bind to GroEL. At that point, the overall machine principle was clear—the nonnative protein would bind in an initially hydrophobic open cavity that would radically reorganize to trap the substrate in a hydrophilic space, perhaps after giving it some forceful tugs Fig. 4). de novo from high-resolution cryo EM maps or deduced from docking of previously known domains, have been arriving in the PDB in increasing numbers, now totaling over 9000 entries. During this development, the PDB and the field of protein crystallography in particular have provided a role model for the developing field of cryo EM. The organization created by the structural biology community with the PDB and the collaborative software project CCP4 (https://www.ccp4.ac.uk/) provided a path for organizing the scientific community, sharing data and software, and developing standards. The EM databank, EMDB, was based on the PDB and hosted at PDBe (https://www.ebi.ac.uk/pdbe/).In recent years, the focus for structural biologists interested in large macromolecular machines has shifted to cryo EM, as multiple advances combined to fuel an explosion of progress. As the resolution of single particle EM has improved, atomic models, built either The 1990s saw the initial development of the EMDB with a European Union grant awarded to Carazo and Fuller . The curhttps://www.ccpem.ac.uk/) to help with software support and training, and together with the databases, to establish data and validation standards.The early attempts by the EM community to use structures in the PDB were fraught with difficulties. In EM, the coordinate system puts the origin in a corner of the image or map. Crystallographic density maps place it at the center of the map. The first attempts to combine GroEL atomic coordinates and EM maps were amusing—every time someone tried to fit the coordinates into the map, the two things were so far apart that you couldn’t find them both on the display at the same time. They would literally disappear into a point if you zoomed out enough to get them both in the picture. Crystallographic and EM map formats had to be reunited as the two fields came back together after some decades of separate development, so that EM data could be read by the programs without causing software crashes. This reunification started during the 1990s. Now, the EM community has CCP-EM . A receE and F; ).A–C; . ReA–C; . A diffeFig. 5D; , 31).FigMany areas of structural biology have been closely associated with the database/community developments described above, notably the field of protein synthesis, folding, and chaperones, which together provide the system for proteostasis, in addition to other major biological areas such as virology. The ability to share, reuse, and evaluate structural data is an integral part of the huge progress in structural biology.The authors declare that they have no conflicts of interest with the contents of this article.


Background: Balance and locomotion are two main complex functions, which require intact and efficient neuromuscular and sensory systems, and their proper integration. In many studies the assumption of their dependence is present, and some rehabilitation approaches are based on it. Other papers undermine this assumption. Therefore the aim of this study was to examine the possible dependence between gait and balance in patients with neurological or sensory integration problems, which affected their balance. Methods: 75 patients  participated in the study. They underwent balance assessment on Kistler force plate in two conditions, six tests on a Balance Biodex System and instrumented gait analysis with VICON. The gait and balances parameters and indices, together with entropy and cyclograms were used for the analysis. Spearman correlation, multiple regression, cluster analysis, and discriminant analysis were used as analytical tools. Results: The analysis divided patients into 2 groups with 100% correctly classified cases. Some balance and gait measures are better in the first group, but some others in the second. Conclusions: This finding confirms the hypothesis that there is no direct link between gait and balance deficits. Two main functions which decide about every day quality of life are: locomotion, which enables efficient transfer between various places, and balance, which ensures proper and stable body posture during different tasks. Both functions are complex, and require intact sensory systems , and their proper integration by central nervous system, and efficient control signals, correcting emerging disturbances. Emerging in recent years rehabilitation practice, taking into consideration the above described features of balance and gait, and the concept of a patient as an individual interacting with environment during task performance, encourages the gait retraining in the attempt to improve the balance function of the patient . This apIn some studies this assumption is confirmed by experimental data. In a study performed by Guffey at co-workers on healthy 2 to 4 years old children the spatio-temporal gait parameters and balance abilities were assessed. The results proved that gait parameters explained over 50% of the balance scores, indicating the dependence between the two functions . They anIn contrast to the previous studies some results of the rehabilitation procedures aimed at improving balance and gait are inconclusive. The analysis of papers dealing with efficacy and effectiveness of non-aerobic exercise program for patients suffering from traumatic brain injuries revealed, that most studies were done on small, heterogeneous sample groups, interventions were not standardized, and outcome data were of poor quality . ChildreSeventy five patients participated in the study, 52 with diagnoses of neurological diseases , and 23 with sensory integration problems, affecting mainly balance abilities. The group consisted of 38 boys and 37 girls aged from 5 to 17 years old. BMI index was within normal range (taking into consideration age and sex) in all but one patient, who was over weighted. The study was approved by the Local Bioethical Committee, parents of all children and adolescents gave their informed consent, as well as all participant who were 16 or older.Balance assessment was performed using two types of equipment: Kistler force plates and a Biodex Balance System (BBS).The balance was evaluated in two conditions: with eyes open and with eyes closed. In both conditions the patient was asked to stand still, with both feet parallel to each other, with distance between them approximately equal to the patient’s pelvis width. The arms were hanging freely, and during the standing with eyes open he/she was asked to look straight ahead, and with eyes closed to keep similar position of the head. The data was collected at the sampling frequency of 1000 Hz, and the trials’ time was 30 sec. During analysis the data were resampled to 60 Hz. The following parameters were calculated from the data: maximal radius of the sway, average radius of the sway, maximal forward sway, maximal backward sway, maximal sway to the left, maximal sway to the right, total length of the sway path. The number of parameters exceeding the normal values  were als-On a stable platform with eyes open;-On an unstable platform with eyes open (level of stability 4);-On a slightly unstable platform with eyes open (level of stability 8);-On a platform with changing level of stability with eyes open (from level 12 to 4);-Limits of Stability Test (LOS) with moderate difficulty level  on stabl-Modified Clinical Test of Sensory Integration in Balance (m-CTSIB). This test comprised four different conditions: standing on firm surface with eyes open, standing on firm surface with eyes closed, standing on foam with eyes open, and standing on foam with eyes closed.Patients underwent several balance tests on a Biodex Balance System SD. Each test was repeated three times, and each repetition lasted 20 sec. There was 10 sec break between repetitions. The platform’s system calculated a set of indices, and the overall balance index from each test was taken for the further analysis. The patients were situated on the platform according to the requirements described in the BBS manual . The folThe patients underwent also the objective, instrumented gait analysis, which was performed using 12 T40-S camera VICON MX system. The gait analysis was done the same day as the balance tests. The lower body Plug-In-Gait marker set and model were used. The patients walked with their self-selected gait speed, after they got acquainted with the environment, equipment and tasks. Later six trials were captured using the Nexus software. Part of the processing of the captured trials was calculation of the Gait Deviation Index (GDI), which was later averaged for trials and legs. GDI was calculated using the Nexus software. The kinematics from six trials was later averaged in the Polygon software and expressed as per cent of the gait cycle. The averaged data was exported with values representing every 2% of the gait cycle. From the averaged kinematic data the Gait Variables Scores (GVS) were calculated for left and right leg separately . The GaiThe exported averaged kinematic data were also used to calculated the cyclogram index. Cyclogram, called also an angle-angle diagram, reflects the coordination abilities of the patient during gait. On one axis there are values of an angle in one joint throughout the gait cycle, on the second one values of the second joint, both angles in the same plane. According to Goswami cyclograms reflect the kinematic changes within the whole gait cycle and the coupling between the joints . In the All calculations were performed using Statistica software v10.0  and no assumption of the number of groups, and finally the discriminant analysis. Spearman rank correlation test was used to establish the dependence between entropy in eyes open and eyes closed conditions, and to see if there is a link between standard method of balance assessment  and entropy. Multiple regression method was used to check the dependence between gait and balance parameters. Two groups of these analyses were performed: first when entropy and balance indices were dependent variables, and gait indices were independent variables, and second when gait indices were dependent variables, and balance indices independent ones. Cluster analysis was performed to see if patients with neurological and sensory integration problems could be grouped based on their gait and balance parameters. The averaged linkage clustering method was used. In this method the distance between the cases is calculated and the criterion for linking them is the smallest distance. Later the basic clusters are hierarchically link with each other to form the cluster tree. Based on the cluster analysis, i.e. visual inspection of the hierarchical cluster tree, the patients were divided into two groups, and discriminant analysis was used to reveal which parameters were the predictors deciding to which of these two groups a patient was assigned. The division of the patients into two groups was arbitral  were relatively small. As there was no golden standard to which the division can be compared, we used the discriminant analysis as an internal evaluation of the quality of the cluster analysis. Additionally the t-Student test was performed to see if the demographical parameters differ between the two groups. There is statistically significant correlation (moderate) between entropy in eyes closed and eyes open condition: R = 0.469. Other statistically significant correlations found during analysis were in eyes open condition: between entropy and number of parameters exceeding normal values  (R = 0.675), between entropy and total sway path (R = 0.614); in eyes closed condition: between entropy and number of parameters exceeding normal values  (R = 0.683), between entropy and total sway path (R = 0.640). The results of the multiple regression tests are presented in The results of the discriminant analysis, which was done based on this division, are presented in The aim of this study was to examine the possible dependence of the gait pattern and balance problems in patients with neurological problems and balance deficits, as there are contradictory statements in the literature. To better assess if such a dependence exists we used balance parameters from two balance tests performed on the Kistler force plate , and six balance tests performed on the Biodex Balance System. The set of these tests reflected different balance situation, which patients could encountered in the daily life: standing on stable surface, on unstable surface with different stability levels, foam, etc. In case of quiet standing tests performed on Kistler platform apart from most popular parameters used to describe balance ability  the entropy of the sway path was calculated for both tests as a measure of complexity and regularity of the paths. In case of the gait evaluation we used commonly used indices assessing the overall gait patterns such as GDI, GPS and GVS, as well as additional measures: cyclograms and calculated from them cyclogram index, which reflect the inter-joint coordination ability of the patients. The analyses of the data confirmed our preliminary findings ,10 that Multiple regressions revealed some dependences between balance and gait parameters, but in each of these analyses only few parameters were indicated as statistically significant: from one to four. Moreover, there is no consistency between the results: different gait parameters were indicated as statistically significant for different balance parameters and vice versa see , except To check if the conclusion about lack of direct dependence between gait and balance, arising from the results of multiple regression was correct, a cluster analysis of the patients based on the Euclidean distance between them was performed. All gait and balance parameters were used for the clustering  of casesThe summary statistics of all parameters  calculatThe posture control develops in children from birth till the adulthood, with most intensive development till the 6th year of age ,16, and Practically in all creatures rhythmic and stereotyped motor tasks, like locomotion, swimming and flying are controlled to a large extend by spinal central pattern generators, which, in vertebrates, are capable under specific conditions, to produce rhythmic and coordinated movements, even in absence of descending of peripheral inputs . Proper One of the limitations of the study is the unbalance number of patients in the groups which arisen from the cluster analysis. This was not intended: the analysis was performed after the recruitment process was finished and all the data were collected. Maybe the proportion (approximately) 1:3.5 reflects differences between neurological patients concerning gait and balance deficits, but more studies to confirm this finding are needed.


The aims of this review are to detail the immunocryosurgery protocol with the addition of daily practice clinical tips, compile data on the mechanism of action of immunocryosurgery, and delineate indications and possible future applications. Most of the available data originate from the treatment of BCC, of all histological types and localizations, and the principles reported mainly reflect on evidence related to the treatment of this common skin cancer.Immunocryosurgery is a minimally invasive combinational therapeutic procedure that has been designed, developed, and evaluated in the Dermatology Department of the University of Ioannina from 2004. In a fixed time protocol, this approach combines immune stimulatory therapy with imiquimod and cryosurgery, i.e., cryosurgery is applied during continuous imiquimod treatment. Laboratory findings in tissue and blood level credit the efficacy to the synergy of imiquimod and cryosurgery. The synergy has been established through clinical trials and the excellent feasibility and efficacy demonstrated in clinical practice. Immunocryosurgery has extensive proof of excellent efficacy, comparable to surgery, in the treatment of basal cell carcinoma. It has also been evaluated in cases of Bowen’s disease, keratoacanthoma, Merkel cell carcinoma,  Immunocryosurgery is a minimally invasive combinational therapeutic procedure that has been designed, developed, and evaluated in the Department of Dermatology in the University of Ioannina Medical School from 2004 . In a filentigo maligna [The combination of two therapeutic approaches is based on the synergy or at least on the complementarity of the respective therapeutic mechanisms. Thus, during the development period, immunocryosurgery was designed based on general principles of cancer immunotherapy and the theoretical implementation of published data on the anticancer modes of action of the two combined modalities . The pos maligna , and, mo maligna  with or  maligna ,3,11,12.The aims of this review are to detail the immunocryosurgery protocol with the addition of practical tips for its successful application in the every-day clinical practice, compile data on the mechanism of action, and to delineate indications and possible future applications. Most of the available data originate from the treatment of BCC, of all histological types and localizations, and the general principles reported herein mainly reflect on evidence and experience related to the treatment of this common skin cancer.The combination of imiquimod and cryosurgery has a spectrum of synergistic actions that result in the enhancement of their respective therapeutic action ,14. It sThe precryosurgery treatment of the tumor with imiquimod creates a proapoptotic environment within and around the tumor  that intImmunocryosurgery may further enhance vascular tumor bed destruction by promoting initial damage of vessel endothelia  and by restricting the induction of adequate tumor neovascularization after the cryosurgical insult ,17.Finally, data originating from BCC studies suggest that a cascade of tissue events during immunocryosurgery leads to an efficient antitumor immune response .+ T cells in the tumor tissue had significantly already increased two days after the cryosurgery session. At the same time, the initial, during the early, precryosurgical phase of imiquimod treatment, increased infiltration of the tumor by Treg (Foxp3+) cells is reversed. Conclusively, the net effect of cryosurgery during imiquimod treatment of BCC is an abrupt increase in the CD3+/Foxp3+ ratio at the tumor site. This finding suggests that the tissue perturbations created by the cryosurgery session may probably play a decisive modulating role in the cellular composition of the inflammatory infiltrate during immunocryosurgery, eventually inducing an effective tumor-destructing immune response. Analogous synergy of imiquimod and cryosurgery has also been described at a preclinical level against tumors inexperimental mice, i.e., B16-OVA melanoma [Contrary to surgery, which removes the tumor tissue, immunocryosurgery as an ablative therapeutic modality yields tumor destruction in situ, enabling a vivid intervention of immunological mechanism. Tumors have developed diverse mechanisms to defend against host’s immune attack . BCCs, fmelanoma .In conclusion, immunocryosurgery seems to set in to action an in situ mechanism that resembles an antitumor vaccination.2 from the Cryogun. The latter is adapted with a large tip (A or B size) in order to ascertain rapid freezing. The second freeze-thaw cycle of the session is similarly performed as the first after complete thawing of the lesion. According to our experience, monitoring tissue temperature or thaw duration times do not really impact treatment outcome. If required, e.g., for larger BCC, treatment cycles can be repeated as indicated, or sometimes the duration of the treatment can be extended with continuous application of imiquimod for several weeks and repeated sessions of cryosurgery every 2–3 weeks [Immunocryosurgery is applied in typical 5-week cycles (“a standard 5-week immunocryosurgery cycle”). The right timing and scheduling of each treatment modality is essential to optimize the anti-neoplastic effect. The imiquimod cream  is applied every night as a thin layer on the lesion and a 0.5 cm rim of surrounding skin. The cryosurgery session is performed on day 14. It consists of two freeze-thaw cycles. Each of them constitutes of 10–20 s of active freezing of the tumor and a 0.5–1.0 cm-rim around it . Given t–3 weeks .During immunocryosurgery, a strong inflammatory reaction can develop at the treated site. This is usually more prominent in the first week after the cryosurgery session and lasts up to a few days after the end of treatment. Patients can wash the treated area every morning with simple soap and water along with gentle removal of scabs and crusts that build up there. It should be noted that, often at the end of the treatment, a thick, strong, inelastic, steadily adherent crust develops which can then be removed after closed application of an emollient cream for 30–60 min. This crust has been more rarely reported after treatment with imiquimod monotherapy and has been eloquently termed “arbor cutis” .lentigo maligna, keratoacanthoma), immunocryosurgery can be used as an adjuvant or alternative to standard therapeutic modalities, such as surgery. The contraindications of immunocryosurgery do not differ from those applied for each of the two combined modalities, i.e., imiquimod and cryosurgery. It should be stressed that treatment of tumors in patients under iatrogenic immunosuppression, including solid organ transplant recipients, is not a contraindication.Eligible for treatment are all patients with a BCC or Bowen’s diseases that can follow instructions and can apply imiquimod for the defined period of time. For the other diagnoses  is often observed after a successful treatment. Nevertheless, the aesthetic outcome of the treatment is judged as “good” or “excellent” by doctors and patients (own unpublished data). Interestingly, some patients report a tingling sensation on the treated area after more than a year, which however does not interfere or disturb their daily activities ,31.The main advantages of immunocryosurgery from the health system perspective include the low implementation cost, the easy applicability at an outpatient level ,32, and As already mentioned, the main disadvantage of immunocryosurgery is the topical discomfort caused by the induction of the inflammatory response in the treated area. The patients should be encouraged to continue treatment and be reassured for the temporary nature of the irritation. It is usually helpful to demonstrate series of typical cases   in orderSubsequently, we will comment on two issues that commonly arise during treatment and are important to strengthen the doctor–patient trust and to achieve the mandatory compliance to the therapy.In almost all cases an intense inflammatory response develops after the cryosurgery session, even in those patients that after 14 days of imiquimod application presented with minimal inflammation at the tumor site. This is considered a positive predictive sign of therapy outcome, yet there are no means to predict the intensity of this reaction before treatment as it varies significantly between patients but also between lesions within the same patient. Generally, it is more pronounced in older patients with many concurrent actinic keratoses and photodamage . HoweverAbsence of inflammation is an exception to the rule and can be observed in a few cases, particularly in patients with underlying hematological malignancies or under immunosuppresive treatment. In these cases, the desired inflammatory response is achieved with the addition of a topical retinoid (tazarotene or tretinoin) every morning on the treated area.In contrast to surgery where compliance during the treatment procedure is not an issue, the application of imiquimod is totally dependent on the patient and therefore detailing the procedure and the anticipated adverse events is of paramount importance. We have observed that patients that fail to apply imiquimod the previous 1–2 days prior to the cryosurgery session have a reduced clearance rate.After the cryosurgery session, it is not uncommon that the patients, especially those with large lesions treatments, may present with some flu-like symptoms including low-grade fever, anorexia, and tiredness. The patients can be reassured that most symptoms commonly subside 7–10 days after the cryosurgery session or certainly within a week after the end of treatment. Again, demonstrating pictures of the treatment procedure is also reassuring to the patient. Availability of the treating physician for a short telephone call or a visit is also important. It should be stressed that, although during immunocryosurgery some “grotesque” clinical pictures of the treated area may develop , these aImmunocryosurgery is a relatively new ablative addition to the available skin cancer treatments, and the spectrum of indications is still under investigation. Principally, it has been used in the treatment of keratinocytic skin cancers, with more experience focusing on the treatment of BCC. Currently, no studies exist that compare directly immunocryosurgery with the standard of care which is surgery. Published data on the use of immunocryosurgery from us or other authors are summarized in The selection of the suitable non-surgical, ablative treatment of keratinocytic skin cancers is determined by the histological diagnosis as well as the general physical condition of the patient, the co-morbidities, and the concurrent medications. These general rules also apply for immunocryosurgery. In general, immunocryosurgery is indicated for the treatment of BCC, Bowen’s disease, and as field treatment of actinic keratoses. Under certain circumstances, it could be considered as a treatment alternative for actinic cheilitis, keratoacanthoma, and low-grade SCC.The efficacy of immunocryosurgery in these epithelial cancers and precancers is highly dependent on the stage of the tumor and its inherent biological behavior. In principle, immunocryosurgery is highly effective and thus indicated as a treatment of in situ  or low grade (keratoacanthoma) cutaneous neoplasms. However, this cannot be expanded to include cSCC as it has unpredictable behavior which will be further explained in a following section.2) or multiple confluent lesions. However, application of imiquimod in an extended large field of photodamaged skin  can induce extreme inflammation beyond the clinical margins of the lesions [2, of only 10 s each. The patient applies the imiquimod cream the same day of the cryosurgery session and every other night for four subsequent applications. In total, imiquimod is applied for 11 nights, and, to assist the patient, a detailed application calendar can be given. For lesions with a retarded response, this cycle can be repeated and in the absence of inflammation a topical retinoid can be added. This approach can be used also for the treatment of hypertrophic AK.In the treatment of actinic keratoses, immunocryosurgery combines the strong therapeutic potential of a spatially targeted treatment (cryosurgery) and a field therapy (imiquimod). Thus, it is suitable for treatment of large anatomical areas  within a large treated area, cryosurgery monotherapy can suffice while, for larger lesions, a repeat immunocryosurgery cycle will be usually needed. It should be stressed that, up to now, and in contrast to observations reported for imiquimod monotherapy [Both cryosurgery and imiquimod are considered as moderately effective modalities for Bowen’s disease . Howeverotherapy , we haveotherapy ,38.Keratoacanthoma diagnosis entails a certain degree of uncertainty as it cannot always be safely differentiated from cSCC ,49 and, 2, open spray, 2 cycles of 15 s each) followed by 5 mg of intralesional methotrexate immediately afterwards. In the following 2 weeks, the patient applies imiquimod every night on the lesion. The cryosurgery session is repeated in two-weekly intervals until complete regression of the tumor.Regarding immunocryosurgery, we have effectively used this approach in combination with intralesional injections of methotrexate as an alternative to the treatment of keratoacanthoma . After tBy transferring our experience from large BCC ,33 , locally advanced BCC , as well as multiple concurrent or metachronous BCC.To date, the larger part of the experience in the treatment of BCC with immunocryosurgery entails tumors with a maximal diameter up to 2 cm, irrespective of anatomical localization, histologic diagnosis, both primary and relapses after surgery. These cases represent 80–90% of all BCC ,59 and tRegarding the efficacy of the modality, it is comparable to the first-line surgical therapy. Notably, also in cases of no-response or relapse after one cycle of immunocryosurgery with a second standard 5-week cycle, a 99% clearance rate of BCC with a maximal diameter of ≤2 cm is achieved . This reIt should be stressed that the efficacy of immunocryosurgery is significantly higher in comparison to imiquimod monotherapy for nodular BCC or to that of cryosurgery alone, particularly for the employed moderate freezing times <20 s. Notably, tumors that are indicated for Mohs surgery, such as the ones with undefined borders or in localizations prone to relapse after surgery, do not constitute a contraindication to immunocryosurgery.In the recent European Guidelines for the diagnosis and treatment of BCC, immunocryosurgery is recommended as one of the alternative treatments for tumors that are not anticipated to have a satisfactory outcome with surgical excision . Accordi2 overlapping sectors, sometimes under local anesthesia. During follow-up, diagnostic biopsies are performed and, if necessary, immunocryosurgery can be repeated.This corresponds to tumors with a maximal diameter of >2 cm, in which the efficacy of a standard 5-week immunocryosurgery cycle is restricted as the tumor size increases. For tumors 3–5 cm in maximal diameter, the efficacy is confined to approximately 75%through our own empirical observations. For these tumors, we apply longer cryosurgery sessions (2 × 20 s) and the tumor is frozen in 1–2 cmCurrently, we are also evaluating longer treatment cycles  or the aBCC can appear as multiple tumors in patients that have been subjected to carcinogenic and/or immunosuppressive factors . These tumors are prone to confluency which threatens vital anatomical structures or constitute surgical fields of many centimeters where surgical treatment or therapeutic radiation would constitute a challenge as it would leave significant anatomical or functional residua on the treated area.Immunocryosurgery ascertains the maximal preservations of tissues and constitutes an ideal treatment selection for these patients. At this point, a special note pertaining to patients with Gorlin–Goltz syndrome is required. Although we did not have the chance to treat a patient with this diagnosis, taking into consideration the data from the independent efficacy of imiquimod and cryosurgery ,63, we sFor some patients with locally advanced BCC, palliative therapy is a more realistic approach in comparison to cure. This applies for neglected cases of BCC which are not amenable to conventional therapy as they have progressed to aggressive tumors with potential of unlimited local extension. In these cases, immunocryosurgery can be used as a palliative intervention with the aim to reduce the tumor load and protect significant anatomical structures. It would be interesting to enquire combinations of immunocryosurgery with systematic therapies for locally advanced BCC, such as the hedgehog inhibitors  or the recent addition of the immune stimulating anti-PD antibodies . A combiLentigo maligna is an in situ melanoma that has the potential to proceed to an invasive melanoma in 5% of the patients during their lifetime. Both monomodal imiquimod and cryosurgery have been used in the treatment of lentigo maligna [lentigo maligna [lentigo maligna [lentigo maligna; this can be achieved with repeated cryosurgery sessions during continuing imiquimod application, as required [ maligna ,65 as we maligna . Additio maligna , includi maligna . As withrequired . Close fTo date, immunocryosurgery is not recommended for the treatment of malignant melanoma. However, there are two instances in which it could be used as an alternative or adjuvant treatment: (i) After a diagnostic excision biopsy in order to treat possible remnants of the tumor in anatomical areas where a wider excision according to the guidelines could compromise vital structures. In contrast, in immunocryosurgery the inflammation induced is temporary and the therapy only targets the involved by the tumor area, as the experience from the treatment of BCC shows. (ii) As palliative therapy for cutaneous melanoma metastases. In the latter case, the induction and maintenance of a considerable inflammatory response is of paramount importance; for this reason, extended treatment schemes with repeated cryosurgery sessions and/or adjuvant topical retinoids could be used until clinical regression is achieved ,67.2 area each time. The patient starts to apply imiquimod in other lesions at the end of a 5-week treatment cycle. Interestingly, the induced inflammatory response is not that intense in comparison with the treatment of keratinocytic skin cancers, although the efficacy is satisfactory when we compare the treated with the untreated areas.Skin lesions of classic sarcoma Kaposi respond promptly to immunocryosurgery. Until now, we have applied this approach to elderly patients that were concurrently treated with interferon-α . The lesMerkel cell carcinoma is a rare aggressive malignant tumor which appears in elderly patients and is characterized by local relapses after surgical removal of the primary tumor. We have described the treatment of in transit lesions appearing in a patient 6 months after the removal of the primary carcinoma . In partImiquimod is licensed for the treatment of genital warts and cryotherapy  and is aFor large, outgrowing lesions, we recommend the standard daily scheme and cryosurgery sessions every two weeks according to the clinical response (flattening) of the lesions.Plantar warts can prove therapy-resistant, especially in subpopulations of immunocompromised individuals. The alternate day immunocryosurgery scheme with cryosurgery every two weeks could be used as an efficacious alternative for resistant warts. As a rule, minimal inflammation is observed with this approach; in case of failing or delayed response, the frequency of imiquimod application can be increased to a daily scheme for non-responders.2, 2 cycles, 10 s each) is applied every two weeks guided by the clinical response. Excellent healing without scaring is achieved, and, interestingly, the treatment of these lesions is not associated with distinct inflammation.Imiquimod and cryosurgery as monotherapies are used for the treatment of pyogenic granuloma ,70. We hIt should be stressed that the diagnosis of pyogenic granuloma should be confirmed in order to avoid the delay of treatment of an amelanotic melanoma .In conclusion, the combination of imiquimod and cryosurgery within the frame of immunocryosurgery is a potent and efficacious treatment for keratinocytic skin cancers that enables a lasting therapeutic result combined with maximal healthy tissue preservation. BCC remains the main domain of immunocryosurgery application. However, the spectrum of diagnoses that are amenable to treatment with immunocryosurgery is currently actively explored, as well as the impact of possible adjuvants.


A growing number of epidemiological evidence suggests a significant increase in waterpipe tobacco smoking, and its potential to become a major public health concern in most Arabic countries, including Saudi Arabia. A cross-sectional study was carried out to assess the prevalence of intention to quit among ever users of waterpipe and intention to start among the never users. The study also investigated the barriers that may prevent users from quitting or trigger the nonusers to start waterpipe smoking. The study consisted of 464 university students from Eastern Province, Saudi Arabia. One hundred and sixty-eight (36.2%) participants were responded that they had WTS at least one time in the past. Among the ever users of WTS, 120 (71.4%) participants had made an attempt or more to quit WTS in the past, 64 (38.1%) had made more than one attempt, and nearly two-third expressed the intention to quit WTS in the future. Forty (13.5%) out of 296 never-users expressed their intention to start WTS in the future. The study further showed that peer influence, social acceptance, and risk perception were significant predictors of intention to start or stop WTS among students.  It is promising that substantial users have the intention to discontinue WTS, though a fraction of never users wish to try WTS in the future. The water pipe device indirectly heats tobacco and produces smoke, which then passes through a column of water before being inhaled through the mouth using a pipe. It has different names, such as waterpipe, hookah, narghile, and arghile. A growing number of epidemiological evidence suggests a significant increase in waterpipe tobacco smoking (WTS), and its potential to become a major public health concern in most Arabic countries, including Saudi Arabia . The increased use of WTS among the Saudi population may be attributed to different reasons, including cultural reasons and the misconception that smoking waterpipe is safe and not as harmful as smoking cigarettes. According to the World Health Organization (WHO) and other studies, WTS may be addictive as in other forms of tobacco use and may cause similar health risks that smoking cigarettes can cause, including lung cancer, respiratory illness, and low birth weight .As part of tobacco-free initiatives, WHO has introduced MPOWER measures to promote government action on tobacco control . MPOWER consists of six evidence-informed and cost-effective interventions: Monitoring tobacco use, Protecting people from tobacco use through smoke-free policies, Offering cessation programs, health Warnings, Enforcing a ban on promotion, and make tobacco products less affordable by Raising taxes on them. In line with these measures, Saudi Arabia has enforced laws preventing smoking in most public places . The country further ensured the availability of smoking cessation support through primary care health centers, availability of nicotine replacement therapy, Bupropion, and Varenicline through pharmacies with full health insurance coverage, and offered a toll-free telephone help-line to discuss cessation . In the country, health warnings on tobacco packages are mandated by law, and a number of anti-tobacco mass media campaigns were aired . As per the WHO report on global tobacco pandemic, Saudi Arabia has recently implemented the MPOWER measures at the moderate to best practice level except on the monitoring aspect . A significant reduction in the prevalence of tobacco use would be expected if the implementation of these measures at the highest level . The overall reduction is expected through the increase in smoking quit rate and reduction in smoking initiation rate.The smoking-related beliefs and the intention to quit or start smoking among Saudi youth have been the focus of smoking cessation research. The smoking-related beliefs could be explained by a health belief model that is used to predict an action that people might take based on the belief to prevent, to screen for, or to control disease situations . The first component of this model is the perception of susceptibility which refers to beliefs of users regarding the likelihood of having some negative consequences of this habit . The second component is the perception of the seriousness of the health outcomes and the possible social of smoking that might impact the decision of people toward quitting or starting smoking . The third component of the model is the perception of the benefits of quitting or starting smoking. Furthermore, the perceptions of barriers and obstacles such as social barriers  could lead the person to decide either to start or to quit waterpipe smoking . There is a lack of studies to investigate the factors related to intention to quit or start waterpipe smoking among smokers in Saudi Arabia. The present study has two objectives: 1) to determine the prevalence of intention to quit among the WTS users and the intention to start among the nonusers; 2) to examine socio-demographic characteristics, smoking-related beliefs as predictors of intention to quit and intention to start WTS among Saudi university students. The impact of our study was that it might help to know in-depth the social, cultural, and other factors around smoking waterpipe, in order to overcome all the barriers that may prevent users from quitting or trigger nonusers to start WTS.Study design and sampleWe conducted a questionnaire-based cross-sectional study among university students. The study sample was selected from various colleges of a leading public university in the Eastern province of Saudi Arabia. The university has more than 45,000 registered students over 21 colleges that spread over the Eastern Province, Saudi Arabia. The study sample comprised of male and female undergraduate students aged below 18 years under health, engineering, and arts and science stream of studies. A minimum sample size of 323 participants was required to estimate a prevalence of 30% with 5% absolute precision and a 95% confidence level. Therefore we targetted a sample of 500 students with the expectation of incomplete data from some students. A group of final-year students had been selected and trained for data collection using a questionnaire. The data collectors approached students at public places, such as the atrium, cafeteria, library, and other open areas, within the university in order to collect the data. The purpose of the study was explained, and consent for participation was obtained from the participating students. The participants were requested to fill the questionnaire with minimal support from the data collectors. Ethical approval was obtained from the Institutional Ethics Committee, College of Clinical Pharmacy, Imam Abdulrahman Bin Faisal University.QuestionnaireThe questionnaire has four sections: socio-demographic information of participants, smoking status and family history, the perceptions on WTS, and the information on the willingness to quit or try WTS in the future. The first three sections were adapted from literature on WTS use .Socio-demographic detailsData on age, gender, study stream, marital status, area of residence , and monthly family income of participants were collected.Smoking status and family historyParticipants were asked if they had smoked cigarettes, e-cigarettes, and waterpipe in the past ever in addition to a question on their 30-days use. The frequency of WTS use among friends and family members were also obtained.Perceptions on WTSThe questionnaire contained eight items to assess risk-perception about WTS, knowledge on the hazards of WTS, and the normative belief on the social acceptance of WTS. The beliefs about the addictive and harmful effects of WTS compared to cigarettes were rated on a five-point scale. The lowest score of ‘0’ indicated ‘WTS is much less addictive/harmful than cigarettes’, and the highest score of 4 indicated ‘WTS is much more addictive/harmful than cigarettes’. The overall risk-perception was assessed using the sum of the two items with a lower total score indicates the higher the false perception. The knowledge on the hazards of WTS was assessed by asking the participants to rate five statements about WTS. The statements were 1) water filters the toxic elements of WTS, 2) WTS is free of tar, 3) WTS is free of nicotine, 4) WTS is free of carbon monoxide, and 5) no increased risk of cardiovascular diseases (CVD) due to WTS. The participants rated the statements on a five-point Likert scale ranging ‘strongly agree (score 0)’ to ‘strongly disagree (score 4)’. A total score of these five items were calculated as a measure of knowledge on hazards of WTS with a lower score indicates lower the knowledge. In order to assess the normative belief on social acceptance of WTS, participants were requested to rate their belief on acceptance of WTS in the society on a four-point scale ranging ‘not acceptable’ to ‘very acceptable’.Intention to quit or start in the futureParticipants who had used WTS in the past were asked about their previous attempt to quit, previous use of cessation mediations/programs, and their willingness to quit in the future. Their self-confidence with the quit decision was also rated. Participants who never used WTS in the past were also asked their intention to try WTS in the future.Data analysis‘Ever user’ of WTS was a participant who used WTS in the past, while ‘never user’ of WTS was a participant who never used WTS in the past. We estimated the prevalence of intention to quit among the ever users and intention to try among never users. For the purpose of analysis, the total risk perception score and knowledge score were categorized into three levels using tertiles: categories with high scores (low favored for WTS), moderate scores, and low scores (highly favored for WTS). Further, categories of individual items were collapsed into three categories. Data were summarized using frequencies and percentages. A Chi-square test for association was used to identify the predictors of intention to quit and start WTS in the future. A p-value of less than 0.05 was considered statistically significant. Analyses were carried out using SPSS Statistics version 24.0.The study sample consisted of 464 participants, of which 268 (57.8%) were males. The description of the participants was detailed in Ever users of WTSOne hundred and sixty-eight (36.2%) participants were responded that they had WTS at least one time in the past . Column Intention to start WTS in the future among never-usersThere were 40 13.5%) out of 296 never-users were expressed their intention to start WTS in the future . The pro.5% out oPrevious quitting attempt and intention to quit WTS in the future among ever usersAmong the ever users of WTS, 120 (71.4%) participants had made an attempt or more to quit WTS in the past; 64 (38.1%) had made more than one attempt. More than half of users (94/168) were aware of cessation services for WTS. However, only 14.3% (24/168) had used the service: Bupropion was used by four participants, Varenicline or NicoDerm CQ was used by 16 participants, and other medications were used by another four participants.Among the ever users of WTS, nearly two-thirds (106/168) expressed the intention to quit WTS in the future . The proThe present study investigated the prevalence of the previous attempt to quit, and intention quit WTS among waterpipe ever users and the prevalence of intention to start WTS among the never users in the Eastern Province, Saudi Arabia. Further to aid in developing effective strategies for discouraging the use of WTS among youngsters, the study further investigated the differences in their characteristics, including socio-demographics, knowledge, and perception about WTS, between users with and without intention to quit WTS and nonusers with and without intention to start WTS. The study population was undergraduate students. The study shows that more than one-third (36%) of participants were used WTS at least once in the past with a higher proportion among males than in females. A previous publication was reported that the past 30-day prevalence was 23% with a narrow gender gap . The previous attempt or the intention to quit may explain the gender difference in proportion. Availability, affordability, and attractiveness of waterpipe may be the key reasons for the higher prevalence of WTS use among youngsters . We found that 63% of ever users expressed the intention to quit, and 14% of never users expressed their intention to start in the future. A study from Qatar reported that more than half of waterpipe users admitted to intending to quit the use . A similar proportion of intention to quit smoking was reported among cigarette smokers in Saudi Arabia . Although it is promising that substantial users have the intention to discontinue WTS, still a number of never-users expressed their intention to try WTS in the future. Thus, it is quite important to introduce a more pervasive restriction on WTS in public places and make it less attractive in order to help the students to refrain from WTS. Health warning labels are found to be effective in encouraging students to quit WTS .Previous studies have expressed concern over awareness about WTS cessation techniques among the general public and particularly, among the practicing physicians . The present study reported that nearly three-fourth of ever users had made an attempt or more to quit WTS in the past. Further, only 14% used a cessation service for WTS, though half of WTS users were aware of such cessation services. More importantly, a systematic review highlighted the lack of evidence on the effectiveness of interventions targeting prevention and cessation of WTS and a need for higher quality effective clinical and behavioral interventions .The study found that more males expressed the intention to quit WTS among the users, and in contradictory, more males also expressed the intention start to start WTS in the future among never users. Thus, the imbalance in the proportions leads to a narrow gender gap on the prevalence of past one-month use of WTS, as reported in our previous study. Our finding was in line with a previous study that assessed the willingness to quit cigarette smoking among youngsters, and the study observed that a substantially higher proportion of males preferred to quit smoking than females . The higher proportions among the male students may be linked to their confidence to stop WTS at any time point.We observed, as reported in Smoking behavior of family members and peers is strongly associated with the initiation of WTS among youngsters and women . A similar association was observed in our study. The study found that WTS use was higher among students with a family history of frequent use of WTS. Importantly, the intent to start rate and the intent to quit rate was higher among the students had the family history of use and did not have the family history, respectively. The smoking behavior among the closed ones may contribute to the level of perceived social acceptance of WTS. Among the never users, only 7% of students who think WTS is not socially acceptable were showed intention to start WTS while it was 23% among the students who think WTS is well accepted. Among the ever users, 92% of students who think WTS is not socially acceptable were showed intention to quit WTS while it was only 30% among the students who think WTS is well accepted. This indicates the perception of social acceptance of WTS has a significant effect on preventing the participants for refraining from WTS. Therefore, appropriate measures need to be taken for discouraging the use of waterpipe during social gatherings in order to prevent from perceiving WTS as a kind of entertainment and is considered acceptable in society.Previous studies have demonstrated that the higher prevalence of WTS use is associated with a high level of perceived lack of harmful and addictive effects of WTS among university students . In the current study, more than one-third of participants were misconceived that waterpipe is less harmful and less addictive than cigarettes, and a higher proportion of them were WTS users. We observed that the willingness to quit among the users is significantly associated with the misconception. In overall, three-quarters of users with low-risk perception were expressed willingness to quit while only 43% intended to quit among the students with high-risk perception. Among the nonusers of WTS, a similar difference in the proportion of students who plan to initiate WTS between the levels of risk perception, though it was not statistically significant. The findings indicate that targeting beliefs on the harmfulness and addictiveness may influence the willingness to quit among users and desire to try among nonusers. Mandated waterpipe tobacco package warning may result an increase of awareness of the health effects of WTS , and thus increase in the level of risk perception .It was found that students who think WTS is free of tar, nicotine, or carbon monoxide are at higher risk of start using WTS compared to those who think otherwise. Alternatively, the students who think the WTS is free of the hazardous contents are less likely to stop WTS among users compared to those who think otherwise. The users who think that WTS has increased risk of CVD are more prone to quit compared to the users who think otherwise. In overall, though not statistically significant, the desire to try WTS was reported by 18% of the students with knowledge score that highly favored for WTS in comparison with 10% among the students with knowledge score that less favored for WTS. Similarly, 72% of students with knowledge scores that less favored for WTS were showed intention to quit WTS in comparison with 54% among the students with knowledge scores that highly favored for WTS. Thus, it is important to educate and increase the awareness of students and public on the impact of WTS on health. The textual and pictorial warning on waterpipe tobacco boxes is found to be effective in increasing motivation and intention to stop WTS .This is the first study in the Kingdom of Saudi Arabia to investigate the willingness to quit among WTS users and desire to try WTS among nonusers and its association with the risk perception and knowledge on the health effects of WTS. This was a cross-sectional study, and thus causal association could not be established. Prospective studies are required to show how risk perception and knowledge on the health effects of WTS influence the cessation among WTS users and desire to try WTS among nonusers. Though the study sample selection from different colleges ensured participation from the diverse socio-economic background, the majority of participants were from the Eastern Province, Saudi Arabia. In conclusion, the study identified the impact of peer influence, social acceptance, and risk perception on behavioral intention to start or stop WTS among students. It is promising that substantial users have the intention to discontinue WTS, though a fraction of never users wish to try WTS in the future. Further actions such as introducing more pervasive restrictions on WTS in public places, make it less attractive, mandating health warning on waterpipe tobacco packages, and awareness campaign on the effectiveness of interventions targeting prevention and cessation of WTS is warranted in order to help the students to refrain from WTS. DA conceived the idea, and later developed as a study in consultation with RJ. DA and RJ equally contributed to the study design. DA was responsible for the data collection. RJ was responsible for the data management and statistical analyses. DA drafted the manuscript. Both authors contributed to further revisions to the draft. All authors have read and approved the final manuscript.


In recent years, post-approval changes (PACs) for medicinal products have increased faster than the national regulatory agencies can attend to without causing any negative impact. This study presents a proposal for regulatory management based on our analysis of the data available from the national regulatory agencies of Latin America on the total post-approval changes evaluated, and the time spent in the process. A retrospective search on the official websites of competent national regulatory authorities (NRAs) of 14 Latin American countries  was conducted to collect data on post-approval changes in the last 4–6 years, up to January 2021. The NRAs considered were Brazil, México, Colombia, and Costa Rica. Our analysis was focused on the post-approval changes that required approval before implementation, those that were submitted, and those that were submitted and approved for small molecules, biologics, and biotechnological products. The results indicated differences in the regulatory processes and procedures applied by the different agencies. We also found that the implementation of the PACs was directly impacted by limited resources, which puts the medication supply for chronic treatments at risk resulting in serious consequences for patients. For local decision-making, Latin American NRAs should implement regulatory pathways already made by regulatory agencies included in the World Health Organization Listed Authorities on PAC approval to optimize their resources and to ensure the continuity of medicine supply for their patients. Chemistry, Manufacturing, and Controls (CMC) changes in medicinal products are inevitable regardless of their type, category, or characteristics. They can be observed in the form of a technological transfer to the final manufacturer of the products during the developmental phase, changing technical needs due to new findings during the product lifecycle, and continuous improvement in the manufacturing processes and product characteristics .Thus, regulations demand a careful evaluation of all changes and proper follow up in the context of regulatory pathways, regardless of whether it is a drug under investigation or a commercial product. To guarantee the quality, safety, and efficacy of the product, leveraging both product and process knowledge as well as the use of a risk-based approach should allow sponsors to achieve the best path for post-approval change (PAC) implementation and regulators to optimize resources through accelerated regulatory pathways .In a recently published report by the Pan American Health Organization (PAHO) , it was The WHO, which finds and fosters the best capabilities of NRAs to promote the standardization concept and its principles around the world, has raised the need for reviewing the classification level of regulatory agencies. For this reason, it issued a robust and unique version of the “WHO Global Benchmarking Tool” for the evaluation of the national regulatory systems of medical products . It alsoThe designation of a regulatory authority as a WLA is ultimately meant to promote access to the supply of safe, effective, and quality medical products by facilitating reliance on the work and decisions of trusted agencies in the regulatory decision-making process to reduce the extra work and wastage of limited financial resources.In this context, an NRA receives the classification Level 4 (this is the NRA with regulatory systems operating at an advanced level of performance and continuous improvement) , if its For its part, PAHO  had prevBy considering risk quality management an essential part of good manufacturing practice along with other related guidelines, this proposal aims to consider the possible mechanisms that can be implemented among Latin American countries. Our primary purpose is to provide recommendations for the more efficient management of the PACs and ensure the planned supply flow of pharmaceutical , biological, and biotechnological products in the NRAs of less mature countries. Specifically, for this proposal, statistical data related to PACs and posted on the NRAs websites of four Latin American countries , were extracted and analyzed. One of them is an active member of the International Council for Standardization of Technical Requirements for Pharmaceuticals for Human Use (ICH) Management Committee (Brazil), and two others are ICH Observers (México and Colombia). This analysis was supported by the regulatory framework and the criteria for classification of PACs, in the countries of the Stringent Regulatory Agencies, based on risk quality management.For several years, some of Latin America's NRAs have maintained dynamic control and improved their processes, performing routine data collection and publishing reports related to the volume of submissions and ongoing internal processes approved, delivered, or rejected, which involves new registrations and the PACs.However, not all countries in the region have available data on their official websites. There were discrepancies found among health agencies in how the data are registered as well as in their reporting periods because some authorities report for a period of 3–4 years, while others report the data every year; this poses a challenge that should be overcome when conducting an objective analysis.Under this scenario, 4  out of 14 agencies were selected, with the available data related to PACs.To encourage Latin American NRAs to optimize their resources and ensure the continuity of medicine supply for their patients, we also aimed to implement regulatory pathways such as the recognition of local decision-making by the regulatory agencies already included in the WLA concerning the issue of PAC approval. Our approach is based on QRM applied by the manufacturer, confirmed during the good manufacturing practices (GMP) inspection under strict compliance with the guidelines of the WHO and the ICH, which guarantees efficacy, safety, and supply of medicine.To perform this analysis, the official websites of competent NRAs from 14 Latin American countries  were systematically reviewed, and a retrospective search of available data related to PACs evaluated or submitted during the last 4–6 years until January 2021 was done. After reviewing and validating the data with a Sanofi Regulatory team from each country and experts searching for information on the websites, the agencies to be included in this analysis were, specifically, ANVISA-Brazil, COFEPRIS-México, INVIMA, and Ministry of Health-Costa Rica.It is important to mention that the Caribbean Islands were not included in this search of data reported.Since common criteria in the available data collected from these regulatory agencies was not found (neither in the definition of the type of product nor in PAC classification), the analysis focused on:PACs that require approval before implementationPACs submittedPACs submitted and approvedSynthetic, biologic, and biotechnological productsIn this sense, the detailed information used for the analysis is as follows:Classification of PAC: Minor and MajorType of products (ANVISA definition): Synthetic, Generics, Technology-Biologics, Biologics*.Data available:  Total PAPeriod: 2016-Jan 2021*This period is the last period updated available in the ANVISA website up until January 2021.The data available was collected only from 2011 until 2016 since there was no data reported after 2016 . That isType of product (COFEPRIS definition): IV , VI .Data available: PACs approved are reported annually.Target Time for evaluation: 45 days .There was no data on PACs submitted for public use as evaluated and approved in Colombia.Thus, our analysis was based on the Ministry of Health and Social Protection report involving PACs evaluated by INVIMA from 2000 until 2020.Data used: PAC submission data extracted from the Ministry of Health and Social Protection report .Type of product: Not definedPeriod: 2000–2020Type of products: Small molecules and biologicsData available: PACs submitted per yearPeriod: 2017–2020Target Time for evaluation:  Small moFor each country involved, the data analyzed was the following:Total number of PACs processed or submitted during the study period, according to the data collected by each regulatory agency.Average time of assessment after submission annually. We considered a scenario of uniform distribution of the PACs between the years of data collection (considering months consisting of 20 working days) to calculate the number of months required by the regulatory authority to complete the assessment before implementation of the PAC.The focus of this analysis was the calculation of the total number of PACs evaluated per year and the time (in days and months) spent by each NRA to process the PACs. Our analysis also evaluated how the current PAC process impacted the implementation of continuous improvement required to guarantee the quality, safety, and efficacy of pharmaceutical products and their supply. With the exception of ANVISA, all countries included in this analysis did not differentiate between PACs Ia/Ib or lower and PACs type II or higher (according to EMA and FDA classification). There was also a delay in implementing PACs that only require notification in the country of manufacture until emission and reception of approval from the regulatory authorities of the Latin American country where it is commercialized.Our analysis is complemented by the assessment of publicly available regulations and guidelines regarding the requirements to guarantee the products' quality, efficacy, and safety as a fundamental support for regulatory decision-making.Period of Analysis: January 2016–January 2021 ANVISA started data collection in 2016 by classifying PACs as minor and major for synthetic , generic, and biological products, similarly to the EMA and FDA classifications. ANVISA has registered a particular classification specifically for biological products, named “technologic biologic products” (Biotechnology) which has been quantified as a major PAC category for this analysis.Until January 2021, a total of 47.182 PACs was evaluated (most of them generic products), with a time of 2,953 days invested by the regulatory agency team. Please see When the time spent assessing major PACs per category of the product is analyzed, 15% was spent on major synthetic products   while 17% was spent on technologic biologic products (Biotechnology) . Please see Considering that a month has 20 workdays, approximately 47.85 months is necessary for assessing major PACs, distributed across 22.2 months for major PACs of synthetic products and 25.65 months for technologic biologic products.Period of Analysis: 2014–2016 Data collected was related to products classified in categories IV and VI, with Category IV products approved to be sold only with a medical prescription (Rx) and Category VI products approved to be sold Over the Counter.A total of 1272 PACs was approved during these years (~424 per year), with a significant percentage of PACs being approved related to Category IV products used to treat several types of chronic diseases. Even though there is little information related to the time spent for approval or on the kind of products included in group IV, neither data was available. Concerning the submissions processed after 2016, we can infer that the number of PACs submitted has increased over the years considering the new marketing authorization issued by COFEPRIS as well as the continuous improvement of processes during the lifecycle of sold products.It is important to highlight that AMIIF (a Mexican trade association)  reportedPeriod of Analysis: 2000–2020The Ministry of Health and Social Protection  published a final report on the evaluation of the regulation reviewed, with regards to Decree 677 published in 1995 .This report documents the results obtained after assessment by the Ministry of Health, INVIMA, and the World Bank on the current regulations. The results indicate that a total of 36,319 submissions  were processed during the analyzed period, with new products comprising 62.7%  of the total. This report also highlights an increase in PAC's submission throughout the years, which overloaded the INVIMA and impacted the availability of products, as we can see in Period of Analysis: 2017–2020 Reglamento Técnico Centro América). According to the RTCA 11.03.64:11, PACs are classified into two types: one is related to the changes that require approval from the NRA before implementation , while the other is related to changes that only require the NRA to be notified concerning their implementation, which comprises primarily minor issues like the change of material/dimensions of the secondary packaging, change of the primary and secondary package label design, discontinuations of registered presentations, changes in the product safety information, and the change or broadening of the distributors. Please see Costa Rica is a member of the Regulation Technical Committee of Central America  was held in Geneva, Switzerland, from 14 to 18 October 2019. Annex 5 on QMS requirements for national inspectorates is defined as follows: .An appropriate infrastructure encompassing the organizational structure, procedures, processes, resources, and systematic actions necessaries to show robust evidence documented to ensure confidence with regards to a product or service to satisfy requirements for quality.A documented change management system should be established to ensure that changes requests are assessed, approved, or rejected; that appropriate resources are allocated; and roles and responsibilities defined. Any change should be documented, communicated to the personnel, and evaluated after implementation, to ensure objectives are met. The change management system should ensure that continuous improvement is undertaken in a timely and effective manner.It is also important to highlight that appropriate quality indicators and methods should be established to monitor and periodically evaluate the inspectorate processes and level of improvement and service (including contracted-out services) to demonstrate that they were performed as planned and they have met the parameters predefined as the Fifty-fourth Report Quality Objectives in WHO Technical Report Series No. 1025 by the Expert Committee on Specifications for Pharmaceutical Preparations . These qQuality management is a wide-ranging concept covering all matters that individually or collectively influence the quality of a product. It is the totality of the arrangements made with the object of ensuring that pharmaceutical products are of the quality required for their intended use. Quality management, therefore, incorporates GMP and other factors, including those outside the scope of this guide, such as product design and development .GMP Is Aimed Primarily at Managing and Minimizing the Risks Inherent in Pharmaceutical Manufacturing to Ensure the Quality, Safety, and Efficacy of Products.The concepts of QA, GMP, QC, and QRM  are inteThe system of QA appropriate to the manufacture of pharmaceutical products should ensure that there is a system for approving changes that may impact product quality. Regular evaluations of the quality of pharmaceutical products should also be conducted to verify the consistency of the process and ensure its continuous improvement; there is also a system for QRM.Changes are an essential part of the lifecycle of the products in constant improvement. That is why the WHO considers the need for manufacturing sites to have an appropriate QMS to ensure the QRM receives the GMP Certificate.According to the ICH Q9 , QRM canICH Q10  mentionsThis guideline demonstrates how an increase in production and process knowledge can contribute to a more precise and accurate understanding of which PACs require regulatory submission and emphasizes the importance of an effective pharmaceutical quality system in the management of changes during the product lifecycle. Such management will eventually reduce unnecessary expenses and time burdens on the industry and regulators. In the meantime, reliable access to high quality medicinal products for patients is assured while continuous improvement is supported. This may result in decreased variability of products and in increased manufacturing efficiency. Implementation of this guideline can also mitigate drug shortages related to manufacturing and quality issues and facilitate the introduction of innovations in manufacturing.ICH has been ensuring access to therapies while guaranteeing, through the guidelines issued, the quality, safety, and efficacy of drugs. They also minimize the risks in each step involved from the development through the clinical investigation, from manufacture until the final product use.Applying ICH Q9, Q10, and Q12  principlStandardizing the classification of reporting categories by creating a “notification” category where it does not already exist is also important. This move would enable regulators to be informed of minor or moderate PACs, as defined under ICH Q12, and avoid delaying the implementation of such PACs by ensuring they are not mistakenly classified as major PACs. It would also ensure appropriate consistency with the way PACs are managed by the Stringent Regulatory Authorities.In this sense, a common understanding and application of QRM principles could facilitate mutual confidence and promote more consistent decisions among regulators, based on the same information. This collaboration could be important in developing policies and guidelines integrating and supporting QRM practices.It is important to mention that participation in international standardization and convergence initiatives can help to strengthen regulatory systems .A good example of this would be the EMA and FDA because both NRAs have based their regulations on the QRM and adopting PAC classification according to the risk level for health and the impact on the quality, safety, and efficacy of the medicinal products, as well as implementing an annual system of notification regarding modifications classified as minor importance. This process can be implemented without previous approval of what has happened during this period (particularly by the EMA) based on the mutual recognition principle, which establishes that the evaluation of a variation that requires approval from some countries members must be done by one of the NRA involved and that the assessment and decision be adopted by the other regulatory authorities to reduce work duplication , 22.Another important forum is the Pharmaceutical Inspection Co-operation Scheme (PIC/S), which supports regulatory inspections by developing common standards in the field of GMP and ensuring that those standards are consistently implemented across their jurisdictions .The use of reliance  or recogConsidering the mechanisms or agreements established for local regulations, the proposal consists of the following steps:The LATAM NRAs adopt reliance or recognition from the Stringent Regulatory Agencies (SRA) in the respective country of origins.Regional Strategy: once the PAC is approved following the reliance or recognition process by a Level 4 LATAM NRA according to PAHO classifications, the other NRAs in the region could follow suit. Please see The basis of this suggestion is the capacity of the regulatory authority in the country of origin to validate and certify implementation and proper functioning of the effective pharmaceutical quality system (involving the QRM) according to recommendations established in the ICH and WHO Guidelines. This suggestion also considers that COFEPRIS-MÉXICO, ANVISA-Brazil, INVIMA-Colombia, ISPE-Chile, and ANMAT-Argentina are regulatory agencies recognized as NRAr by PAHO in Latin America.The principle of recognition mentioned is supported and described in the REGLAMENT (EC) No 1234/2008 OF THE COMMISSION OF NOV 24th, 2008: Commission REGLAMENT (EC) No 1234/2008 concerns the examination of variations to the terms of marketing authorization for medicinal products for human and veterinary use , to reduReliance promotes a more efficient approach to regulation, thereby improving access to quality-assured, effective, and safe medical products. It can take many forms and can be applied to varying degrees while recognizing or considering assessments, decisions, or authorized information from other authorities and institutions .This proposal can only be implemented if the product approved and sold in each Latin American country has the same formula, manufacturing process, specifications, and analytical method certified by the person responsible for the product's manufacture.By adapting these elements into the Latin America NRA regulations, resources and energy can be focused on the most important PACs while PACs with minor to moderate potential impact on the product quality, safety and efficacy can be processed much quicker.We confirmed the differences in the regulatory processes and procedures among the NRAs through the data collected from the NRAs in Brazil, Colombia, México, and Costa Rica. We also looked at how limited resources directly impact the implementation of the PACs, which puts the medication supply for chronic treatments at risk and subsequently results in serious consequences for patients.Benefits obtained through the implementation of these proposals:Patient Benefits:° Availability of therapies, and a lower risk of back orders which ensures a continuous supply of crucial medicines.° Facilitation of access to innovative therapies.° Treatments with compliant quality, safety, and efficacy within established parameters.LATAM Reference regulatory agency Benefits:° Process optimization.° Improvement of timeline approval and implementation times.° Updated real-time dossiers aligned with the manufacturing country.° Increased of technical and scientific capabilities° Standardization of criteria related to PACs with Stringent Regulatory Agency.LATAM NRAs Agency Benefits:° Optimization of resources used in other areas of interest such as anti-counterfeiting and pharmacovigilance.° Updated real-time dossier aligned with the manufacturer and the NRA used as reference.° Quality dossier standardization among the countries.° Standardization of criteria related to PACs around the region.Health System Benefits:° Reduction in shortage of critical therapies .° Timely implementation of improved therapies with high quality, efficacy, and safety.Pharmaceutical Industry benefits.° Manufacture unification by product.° Reduction of time needed to implement changes required to improve the quality, safety, and efficacy of the product.° Ensure timely supply of the product for each country and reduces shortages and discontinuation of treatment for patients.In this analysis, we systematically reviewed all available NRA official websites of LATAM countries. A potential limitation of this analysis is the availability of PAC data from all NRA websites. At the same time, some discrepancies among countries were found in the data collected, which requires more attention from NRAs in the region.We observed clear differences in the regulatory frameworks of different NRAs in Latin American countries vs. the agencies included in the WLA (like the EMA and the FDA) with regards to PAC classification. PACs classified as a minor variation or Type Ia and Ib (by the FDA and EMA) can be implemented immediately or after 30 days, respectively, by the manufacturer. However, the situation is different in Latin America because once PACs are approved by stringent agencies, some NRAs must wait for approval or authorization which impacts the manufacturers, and distribution centers, as well as causing other supply issues.Therefore, globally standardized and consistent regulatory approaches to PACs as proposed in the WHO's guidance on variations , along wDespite discrepancies in the data available, the analysis shows that the time spent by NRAs per year to evaluate PACs should be considered a key performance indicator to evaluate NRAs efficiency in terms of resource management. They can define a target timeline for approval (not exceeding 6 months) without a negative impact on the improvements required to guarantee the quality, safety, and efficacy of pharmaceutical products and their supply in the region.Some recommendations proposed by PAHO in the current report “REGULATORY SYSTEM STRENGTHENING IN THE AMERICAS”  include:“The NRAs need to prioritize regulatory life cycle management of products, finding ways to better handling them and improving regulatory oversight using a holistic view of the entire life cycle of the authorization, improving the allocation of technical and human resources, and adopting electronic tools to improve efficiency.To implement procedures that enable the use of reliance.To improve publicly available regulatory information as part of good regulatory practices.To take advantage of available tools on GMP information. Make better use of public databases, such as EudraGMDP and WHO prequalification databases, to check the GMP status of individual manufacturing sites.Trading of integration mechanisms can facilitate regulatory strengthening.To develop legal and organizational frameworks.”Agreements signed among NRAs of Latin American countries and NRAs should be considered as the main element of any legal frameworks.The common mission between the Industry and NRAs is to ensure that the available therapies should satisfy the patient's needs on time with optimal levels of quality, safety, and efficacy.It is crucial to emphasize the importance of adjusting regulations to optimize the efficiency of the processes related to the PACs of drug , biological, and biotechnological products by allowing the following:the implementation of ICH Q9, Q10, Q12 principles based on QRM and effective Quality Management Systems, so that only the most significant PACs are submitted to regulators for prior approval,the standardization of the reporting classifications of PACs with those of the Stringent Regulatory Authorities, so that the implementation of PACs is consistently and timeously carried out by manufacturers,the setup of defined timelines for reviewing and approving major PACs, not exceeding 6 months, andthe development of reliance and Recognition processes as regulatory pathways.There should also be built-in contingencies for the possibility of fast-tracking as required, not only to face emergencies like the COVID-19 pandemic but also to allow the continuous improvement required to guarantee product quality, safety, and efficacy.There are some agreements between some National Regulatory Agencies in Latin America with SRA, like the one recently signed between EMA and ANVISA, the Confidential Agreement signed between the Americas'NRAr , and theNRAs of Latin America should implement and optimize their digital platforms if possible. Data collection and metrics related to PACs should be evaluated annually to determine the efficiency of the different measures, and to identify the changes and improvements that need to be implemented in processes and procedures.Additionally, the Latin American agencies should also define a target timeline for PAC approval and performance indicators to improve efficiency.This document provides the industry with views on the “Key Principles” documents and gives a holistic vision of what is needed to deliver timely and easily accessible medicinal products.All authors designed the paper, analyzed the data, and wrote the manuscript.The views expressed in this research paper are the independent views of the authors and should not be understood or quoted as being made on behalf of or reflecting the position of their respective companies or any other affiliation.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.