**5. Translational uses of PRP biotechnology clinical relevance of PRPs**

The goal of this section is not to provide an exhaustive overview of current clinical studies but to identify and briefly describe the miscellaneous clinical applications. PRP is considered investigational because currently, there is insufficient evidence to support the use of PRP for all the indications included below.

The use of PRPs has extended to multiple clinical fields and novel applications are emerging to meet varied clinical needs. The increasing use of PRPs in sports medicine, (after withdrawal of restrictions imposed by the IOC International Olympic Committee) and PRP applications in areas such as dentistry and plastic surgery is expected to drive the financial growth of PRPs, that was estimated in the US market as CAGR annual growth rate of 14% from 2009 to 2016 (\$45m in 2009 will reach \$126m in 2016) (http://www.researchviews.com/healthcare/medical/ orthopedicdevices/Viewpoints.asp x?sector=Orthopedic%20Devices&DocID=10728). The diffusion of PRP is attributed to the biosafety of the product due to its autologous origin.

In general, clinical uses of PRP can be categorized in two. First surgical applications, when PRP is used in surgery as an aid to enhance repair, not only of target tissues but PRP also aims to enhance the healing of all the adjacent tissues damaged during the procedure, and secondly conservative uses of PRP, most often involving more than one application and used in outpatient settings. Besides, the use of PRP associated to other medical procedures (laser therapies, etc) offers potential to enhance such procedures. Table 3 and Table 4.

#### **5.1. Treatment of non-healing wounds**

already marketed usually called predicate device. In these cases 510(k) clearance is evaluated

At the European level there is no harmonized regulatory framework for PRP therapies, and each country has its own approach to PRP regulation within the jurisdiction of national authorities. Devices must comply with Class II-a medical device directive 93/42/EEC. Device approval in EU is overseen in each EU country by a governmental body called a Competent Authority. Instead, the surgical use of PRP can be considered as an autologous graft within

Of note, if regulatory requirements for PRP therapies were over-interpreted, unnecessary work derived therein will increase costs and hamper the clinical use of PRP therapies. This hypo‐ thetical situation would be prejudicial for many patients since advancements in PRP science can provide effective treatments for pathologies with substantial social and economic burden.

Currently most insurance plans do not reimburse for PRP treatment due to the lack of data about their efficacy. Interestingly, in the US Category III, code 0232T is used for emerging technologies and applies for nonsurgical uses of PRP. This code allows data collection to be used to document widespread use for FDA approval and potential reimbursement. Besides, this code will allow the AMA (American Medical Association) to track the use of PRP, since codes T are considered experimental they will require pre-authorization for payment. If a physician feels that the patient would benefit from PRP injections, typically as a step to avoid a more costly and invasive procedure preauthorization for PRP reimbursement should be requested. Presentation of cost savings rationale can be the key to successful preauthorization. In general managers are concerned about physician's plans to get injured patients back to

**5. Translational uses of PRP biotechnology clinical relevance of PRPs**

The goal of this section is not to provide an exhaustive overview of current clinical studies but to identify and briefly describe the miscellaneous clinical applications. PRP is considered investigational because currently, there is insufficient evidence to support the use of PRP for

The use of PRPs has extended to multiple clinical fields and novel applications are emerging to meet varied clinical needs. The increasing use of PRPs in sports medicine, (after withdrawal of restrictions imposed by the IOC International Olympic Committee) and PRP applications in areas such as dentistry and plastic surgery is expected to drive the financial growth of PRPs, that was estimated in the US market as CAGR annual growth rate of 14% from 2009 to 2016 (\$45m in 2009 will reach \$126m in 2016) (http://www.researchviews.com/healthcare/medical/

only for substantial equivalency.

16 Biotechnology

*4.2.1. Reimbursement*

productivity or work.

all the indications included below.

Table 1 shows devices and/or kits for PRP preparation

the surgical procedure as regulated by Directive 2004/23/EC.

PRP has been used to treat non-healing wounds for more than 2 decades. In fact, the topical management of chronic leg ulcers was the first clinical application of platelets outside the blood stream with healing purposes. Theoretically PRP or its derivatives are stimulants for nonhealing wounds; the goal is to re-activate healing. The rapid formation of granulation tissue can prevent further deep tissue involvement and associated co-morbidities.

Complex non-healing wounds can have different etiologies including pressure ulcers, diabetes, venous, arterial, or surgical trauma. Impaired wound healing is the major compli‐ cation that results in the development of chronic wounds often leading to amputations as often occurs in the diabetic foot. Components of lower extremity amputations in the diabetic patient include ulceration (85%), faulty wound healing (81%) initial minor trauma (81%), neuropathy (61%), infection (59%) gangrene (55%) and ischemia (46%) (38]. Actually, the wound healing society treatment guidelines for diabetic ulcers advises re-evaluation of the wound and treatment based on failure to reach a 40% reduction of initial wound size by week 4 (39]. Cost for amputations has been estimated to be between \$ 20.000 and \$ 60.000 per case. PRP is deemed useful in this context because it may provide a way to reduce this cost burden to society.

Several RCT have provided data about the efficacy of PRP or PRP derivatives in non-healing wounds. The initial platelet product known as PDWHF (platelet derived wound healing factors) stimulated the formation of granulation tissue in chronic leg ulcers [40]. Later, PDWHF of pooled donors was also examined in the treatment of diabetic ulcers (41]. In general, concomitant pathologies such as diabetes do not hinder the therapeutic effects of PRP, and autologous PRP is effective in the diabetic foot [42-43] or systemic sclerosis [44].

Despite several studies reporting the benefits of PRP in this setting, a recent meta-analysis of ulcer care studies and PRP failed to show any statistically relevant difference favoring PRP for the treatment of chronic wounds. Nevertheless, conclusions are not sound because they are based on 9 RCTs with high or unclear risk of bias [45]. Additional evidences complementary to RCT data can be obtained from practice based medicine and observational studies. These data obtained in a real-world setting provide pragmatic evidences of PRP benefits.

Actually, some PRP manufacturers such as Cytomedix, AutoloGelTM have created wound registries to evaluate the use of PRP and calculate cost savings based on mean treatment times; this is realistic using patients in pretreatment run-in periods as their own controls during standard wound care treatment [46].

The reduction in treatment time should impact clinical and financial decisions. Significant clinical outcomes indicated many previously nonresponsive wounds began actively healing in response to PRP therapy. Cost effectiveness analysis comparing the potential economic benefit of PRP to alternative therapies in treating non-healing diabetic foot ulcers, using an economic model based on peer-reviewed data showed that PRP resulted in improved quality of life and lower cost of care over 5-year period than other treatment modalities for non-healing diabetic ulcers [47].

#### **5.2. Maxillofacial and oral surgery**

The field of PRP gained new impetus at the end of the nineties when maxillofacial surgeons and dentists introduced PRP to augment oral reconstruction procedures. Since then a number of protocols have been developed for different applications. These include socket filling after molar extractions, implant surgery, PRP mixed with bone grafts during osteodistraction, and in the treatment of mandibular tumor resection. Numerous articles describe earlier stabiliza‐ tion of dental implants when PRP is used to enhance the properties of bone grafts, and to modulate the inflammatory status in the surrounding soft tissues.

The utility in several of these procedures seems evident. However, after meta-analyzing 24 studies addressing the use of PRP in the surgical treatment of periodontal diseases, it was concluded that PRP exerts a positive effect only when used with graft materials for the treatment of intrabony defects, but not in guided tissue regeneration. No significant benefit of PRP was found for the treatment of gingival recessions [48].

In sinus augmentation surgery the combination of PRP with autologous bone graft led to increased bone density at 6 months but not at three months [49].

The use of PRP in children is less known because they need little help for healing since young cells and young blood has stronger healing potential and plasticity when compared to adults. However, in the most frequent congenital facial malformation, i.e. cleft lip and palate, the use of PRP is being investigated [50- 53]. The goal here is to reestablish the maxillofacial arch and to close any oro-nasal communication. Besides, PRP is used in the closure of recurrent cleft palate fistulas.

#### **5.3. Plastic surgery and dermatology**

A recent review in plastic surgery including 15 randomized controlled studies and 25 casecontrols showed that the outcomes were favorable in three main PRP indications: wound healing, fat grafting and bone grafting [54].

PRP is used in breast reconstruction associated to fat grafts because it enhances the survival of fat grafts that otherwise had a tendency to be resorbed by the organism. Similarly, the outcome of plastic reconstruction using skin grafts can be improved taking advantage of the pro-survival effects of PRP.

The use of PRP reduced recovery time in facial rejuvenation. In fact the addition of PRP to lipofilling procedures resulted in a significant reduction in the number of days needed to recover before returning to work or restart social activities. Also the aesthetic outcome was significantly better with PRP [55].

Combined treatments as fat graft, laser CO2 and PRP showed clinical benefits in the treatment of atrophic and contractile scars [56].

PRP combined with erbium fractional laser therapy is effective for treating acne scars or acne, at the same time PRP enhances the recovery of laser damaged skin [57].

PRP injected into the scalp is also used to manage androgenetic alopecia [58]. When the efficacy of the interventions was examined in 64 patients, half of the patients showed a clinically meaningful improvement [59]. PRP is also used in hair transplantation [60]. In this situation, PRP shortens the time for hair formation.

## **5.4. Treatment of orthopedic problems**

The reduction in treatment time should impact clinical and financial decisions. Significant clinical outcomes indicated many previously nonresponsive wounds began actively healing in response to PRP therapy. Cost effectiveness analysis comparing the potential economic benefit of PRP to alternative therapies in treating non-healing diabetic foot ulcers, using an economic model based on peer-reviewed data showed that PRP resulted in improved quality of life and lower cost of care over 5-year period than other treatment modalities for non-healing

The field of PRP gained new impetus at the end of the nineties when maxillofacial surgeons and dentists introduced PRP to augment oral reconstruction procedures. Since then a number of protocols have been developed for different applications. These include socket filling after molar extractions, implant surgery, PRP mixed with bone grafts during osteodistraction, and in the treatment of mandibular tumor resection. Numerous articles describe earlier stabiliza‐ tion of dental implants when PRP is used to enhance the properties of bone grafts, and to

The utility in several of these procedures seems evident. However, after meta-analyzing 24 studies addressing the use of PRP in the surgical treatment of periodontal diseases, it was concluded that PRP exerts a positive effect only when used with graft materials for the treatment of intrabony defects, but not in guided tissue regeneration. No significant benefit of

In sinus augmentation surgery the combination of PRP with autologous bone graft led to

The use of PRP in children is less known because they need little help for healing since young cells and young blood has stronger healing potential and plasticity when compared to adults. However, in the most frequent congenital facial malformation, i.e. cleft lip and palate, the use of PRP is being investigated [50- 53]. The goal here is to reestablish the maxillofacial arch and to close any oro-nasal communication. Besides, PRP is used in the closure of recurrent cleft

A recent review in plastic surgery including 15 randomized controlled studies and 25 casecontrols showed that the outcomes were favorable in three main PRP indications: wound

PRP is used in breast reconstruction associated to fat grafts because it enhances the survival of fat grafts that otherwise had a tendency to be resorbed by the organism. Similarly, the outcome of plastic reconstruction using skin grafts can be improved taking advantage of the

The use of PRP reduced recovery time in facial rejuvenation. In fact the addition of PRP to lipofilling procedures resulted in a significant reduction in the number of days needed to

modulate the inflammatory status in the surrounding soft tissues.

PRP was found for the treatment of gingival recessions [48].

increased bone density at 6 months but not at three months [49].

diabetic ulcers [47].

18 Biotechnology

palate fistulas.

**5.3. Plastic surgery and dermatology**

pro-survival effects of PRP.

healing, fat grafting and bone grafting [54].

**5.2. Maxillofacial and oral surgery**

The use of PRP to solve clinical problems in orthopedics has increased with impetus in the past five years the main reason can be the biosafety of the treatment and the fact that actual strategies for management are insufficient. PRP is used in open and arthroscopic surgery and as a conservative treatment for the management of chronic pathologies, most importantly tendinopathies, chondropathies and osteoarthritis. Main properties are attributed to modula‐ tory effects on inflammation and angiogenesis along with reduction in pain [61-63].

Research in orthopedics encompasses a wide range of applications. In sports medicine, physicians are more deeply engaged in conservative management of tendinopathies and muscle injuries

Proposed clinical and surgical applications include spinal-fusion, osteoarthritis [hip and knee), tendinopathy enhancement of healing after ACL reconstruction and muscle strains. The challenge is to show that PRP is superior to the optimal available treatment.

#### *5.4.1. Bone regeneration*

In some applications PRP is used as a coadjuvant associated to autologous or homologous graft and also to bone marrow graft. There is no compelling evidence to demonstrate the efficacy of PRP alone in facilitating the union of long bones for union of tibial osteotomies or pseudoarth‐ rosis or fractures [64). An randomized study involving 21 participants compared PRP+alloge‐ neic graft versus allogeneic bone alone in patients undergoing tibial osteotomy in the medial compartment in patients with OA [65]. There was a significant difference in the proportion of bones that were united after one year in favor of PRP but evidence from a single trial is insufficient to support routine interventions with PRP.

#### *5.4.2. Lumbar fusion and intervertebral disc degeneration*

Posterolateral arthrodesis in lumbar spine surgery was enhanced when PRP was combined with cancellous bone graft as shown by densitometry in a prospective study [66]. However, a randomized clinical study did not show any benefit when PRP is used with autologous bone in mono-segmental posterior lumbar interbody fusion [67].

Intervertebral disc degeneration is also common in orthopedics and current treatments are of limited value to enhance the regenerative process. In fact some studies point out the efficacy of PRP in reversing the degenerative trend of the intervertebral discs based on basic science research [68]. However this application lacks translation in published reports.

#### *5.4.3. Osteoarthritis*

The efficacy of PRP has been most studied in knee osteoarthritis while few clinical data are available about the therapeutic effects in hip OA.

There are several randomized clinical trials comparing the efficacy of PRP treatment with HA administration. Also PRP has been compared with placebo administration. Two recent metaanalysis [69,70] concluded that multiple PRP injections ameliorate pain and improves function and tends to be more effective that HA administration. Patients with lower levels of knee degeneration achieve better results than more advanced knee deterioration. Although less investigated, PRP injections in the hip ameliorate symptomatology and function, however the clinical level of evidence is low and more clinical studies are needed before claiming therapeutic effects in this joint [71].

#### *5.4.4. Tendon pathology*

Most studies in tendon pathology involve guided injections of PRP. Indeed, the development of real-time imaging techniques such ultrasonography enhances the safety and accuracy of PRP delivery during percutaneous management. Consequently, investigation on the efficacy of PRPs for managing chronic tendinopathies has grown in the last years and the quality of the studies has improved considerably. Limitations of observational studies have been overcome by level I and level 2 clinical trials. However due to tendon diversity and function, the various PRP products and the diversity of application protocols, quantitative synthesis and meta-analyses are difficult to perform.

Most frequent upper limb tendinopathies involve the supraspinatus and the medial and lateral epicondyle. In these pathologies, the quality of clinical studies is high/moderate. A quantitative synthesis evaluating the efficacy of PRP as an adjuvant in rotator cuff arthroscopy failed to show any benefit associated to PRP. The only uncertainty in favor of PRP is that it may decrease the proportion of retears, but this needs further confirmation [72].

Ultrasound guided injections and real-time follow-up has fueled the use of PRP in tendino‐ pathies

A recent metaanalysis examining the conservative management of tendinopathy has shown that PRP provide some benefits in pain [61]. Moreover, subgroup analysis showed a modest reduction of pain in epicondylitis [73]. However, major limitations for pooling data and drawing firm conclusions comprise different outcome measurements and follow-up periods.

The most commonly treated tendons in the lower limb were Achilles, the patellar tendon and the plantar fascia. PRP shows potential benefits in these anatomical locations, current eviden‐ ces are encouraging but limited. The need is clear to compare PRP treatment with the most adequate control for each condition.

#### *5.4.5. Muscle injuries*

Intervertebral disc degeneration is also common in orthopedics and current treatments are of limited value to enhance the regenerative process. In fact some studies point out the efficacy of PRP in reversing the degenerative trend of the intervertebral discs based on basic science

The efficacy of PRP has been most studied in knee osteoarthritis while few clinical data are

There are several randomized clinical trials comparing the efficacy of PRP treatment with HA administration. Also PRP has been compared with placebo administration. Two recent metaanalysis [69,70] concluded that multiple PRP injections ameliorate pain and improves function and tends to be more effective that HA administration. Patients with lower levels of knee degeneration achieve better results than more advanced knee deterioration. Although less investigated, PRP injections in the hip ameliorate symptomatology and function, however the clinical level of evidence is low and more clinical studies are needed before claiming

Most studies in tendon pathology involve guided injections of PRP. Indeed, the development of real-time imaging techniques such ultrasonography enhances the safety and accuracy of PRP delivery during percutaneous management. Consequently, investigation on the efficacy of PRPs for managing chronic tendinopathies has grown in the last years and the quality of the studies has improved considerably. Limitations of observational studies have been overcome by level I and level 2 clinical trials. However due to tendon diversity and function, the various PRP products and the diversity of application protocols, quantitative synthesis and

Most frequent upper limb tendinopathies involve the supraspinatus and the medial and lateral epicondyle. In these pathologies, the quality of clinical studies is high/moderate. A quantitative synthesis evaluating the efficacy of PRP as an adjuvant in rotator cuff arthroscopy failed to show any benefit associated to PRP. The only uncertainty in favor of PRP is that it may decrease

Ultrasound guided injections and real-time follow-up has fueled the use of PRP in tendino‐

A recent metaanalysis examining the conservative management of tendinopathy has shown that PRP provide some benefits in pain [61]. Moreover, subgroup analysis showed a modest reduction of pain in epicondylitis [73]. However, major limitations for pooling data and drawing firm conclusions comprise different outcome measurements and follow-up periods.

The most commonly treated tendons in the lower limb were Achilles, the patellar tendon and the plantar fascia. PRP shows potential benefits in these anatomical locations, current eviden‐

the proportion of retears, but this needs further confirmation [72].

research [68]. However this application lacks translation in published reports.

*5.4.3. Osteoarthritis*

20 Biotechnology

available about the therapeutic effects in hip OA.

therapeutic effects in this joint [71].

meta-analyses are difficult to perform.

*5.4.4. Tendon pathology*

pathies

PRP injections are used in professional athletes; the goal is to accelerate muscle healing, and avoid relapses through true muscle regeneration, i.e. absence of scar tissue. Especially in elite athletes the goal is to achieve rapid healing and resume competition faster than with conven‐ tional care. Although case series provide promising results, two recent randomized controlled trials showed divergent results [74,75]. Protocols and PRP formulations were different in both studies, thus further research is warranted.

#### **5.5. Treatment of eye problems**

#### *5.5.1. Macular hole*

Platelet concentrates were used in the 90s as an adjunct to macular hole healing. PRP is an autologous alternative to fibrin glue with much more biological activity conferred by hundreds of GFs and cytokines stored in alpha-granules. The efficacy of platelet concentrates was examined in a multicenter double blind study involving 53 eyes in the experimental group and 57 eyes in the control group. Injection of autologous platelet concentrates during macular hole surgery improved significantly the anatomic success of the intervention but did not influence visual acuity [76].

#### *5.5.2. Dry eye*

Autologous serum administered topically has been used to treat dry eye symptoms because it can improve not only lubrication but also enhance lacrimal production since its composition includes GFs important in this context such as EGF. However, after metaanalysing current studies there were inconsistency in relation to the benefits provided by autologous serum [77].

As an alternative, PRP derivatives mainly the PRP released supernatant or the PRP lysate are being investigated to improve dry eye conditions, administered topically. Alternatively, in severe cases PRP is injected adjacent to the lacrimal gland [78]. Results indicate a significant increase in lacrimal volume and patient's self-reported improvements. PRP also enhanced epithelial status after LASIK but did not affect the recovery of corneal sensitivity.as evaluated in a controlled study in which the contralateral eye was used as control [79].

#### *5.5.3. Corneal ulcers*

Perforated corneal ulcers have been treated with PRP fibrin in a reduced number of patients (N=11=. In all cases the corneal perforation was sealed after 3-5 days of stability of the mem‐ brane in the ulcer [80]. When PRP eyedrops were compared to autologous serum eyedrops in the treatment of persistent epithelial defects PRP was more efficient than serum [81]. After photorefractive keratectomy, the use of PRP enhanced wound healing and reduced haze formation [82]

In a prospective controlled study in acute ocular chemical injury PRP eyedrops as adjunct to standard care was superior to artificial tears, autologous platelet lysate was also effective for the treatment of refractory ocular GVDH in unresponsive patients [83].

#### *5.5.4. Other therapeutic applications*

Some anecdotic uses of PRP in urology for fistula repair [84], in the management of infantile extravasation injury [85], and in gynecology for vaginal prolapse [86-87] have been published.
