2.2.6 Radioprotectors and Chemoprotectors in the Management of Lung Cancer

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2.2.6 Radioprotectors and Chemoprotectors in the Management of Lung Cancer

Ritsuko Komaki, Joe Chang, Zhongxing Liao, James D. Cox, K. A. Mason, and Luka Milas

R. Komaki, MD, Professor of Radiation Oncology, Gloria Lupton Tennison Endowed Professor for Lung Cancer Research J. Chang, MD, PhD; Z. Liao, MD; J. D. Cox; MD, K. A. Mason, MSc; L. Milas, MD, PhD

Department of Radiation Oncology, The University of Texas, M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030, USA

CONTENTS

2.2.6.1 Introduction 123

2.2.6.2 Thiols as Radioprotective Agents 124 2.2.6.2.1 Amifostine: Preclinical Findings 124 2.2.6.2.2 Amifostine: Clinical Studies 126

2.2.6.3 Prostanoids, COX-2, and COX-2 Inhibitors 128 2.2.6.4 Growth Factors and Cytokines 129

2.2.6.5 Pentoxifylline 130

2.2.6.6 Angiotensin Converting Enzyme (ACE)

Inhibitors 130

2.2.6.7 Radioprotective Gene Therapy: Superoxide Dismutase (SOD) 131

2.2.6.8 Concluding Remarks 131 References 132

2.2.6.1 Introduction

Lung cancer is the leading cause of cancer death in most developed countries. Almost one million new cases of lung cancer occur worldwide each year (Jemal et al.

2004), and the prognosis remains poor with an overall survival at 5 years of only 15% (Jemal et al. 2004).

Between 70% and 85% of all cases are histologically classiﬁ ed as non-small cell lung carcinoma (NSCLC), comprised of squamous cell, adenocarcinoma, large cell, or undifferentiated histology, while the remaining belong to small cell histology (Jemal et al. 2004). At the time of diagnosis the majority of patients present with locally advanced disease and many of them have overt metastatic dissemination. Radiation therapy has traditionally been the treatment of choice for locally advanced disease but has provided limited beneﬁ ts both in terms of local tumor control and patient survival, with 2- to 5-year survival commonly not ex- ceeding 10% (National Cancer Institute Cancer

Gov 2002). However, adding cytotoxic drugs to radiotherapy considerably improves treatment outcome, so that the combination of chemotherapy with radiotherapy has currently become a common practice in the treatment of advanced lung cancer. The addition of chemotherapy to radiotherapy has two principal objectives, to increase the chance of local tumor control and to eliminate metastatic disease outside of the radiation ﬁ eld. The former can be achieved by reducing cell burden in tumors undergoing radiotherapy or by interfering with tumor cell radioresistance factors, thereby rendering tumor cells more susceptible to destruction by radiation. Factors which contribute to tumor radioresistance include the failure of tumor cells to undergo cell death after radiation, the cells’

ability to efﬁ ciently repair DNA damage, continued cell proliferation during the course of radiotherapy, cell radioresistance secondary to hypoxia that commonly develops in solid tumors, and the presence in tumor cells of various abnormal molecular structures or dys- regulated processes linked to cellular radioresistance (Milas et al. 2003a).

Addition of induction (neoadjuvant) chemotherapy to radiotherapy results in an increase in median survival time by approximately 4 months, and the overall survival rates at 2 years range from 10% to 15% (NCI 2002; Milas et al. 2003a,b; Dillman et al. 1990;

LeChevalier et al. 1991). These therapeutic gains have been improved by using concurrent chemoradiotherapy, i.e., by administering cytotoxic drugs during the course of radiation treatment (NCI 2002; Milas et al. 2003a; Komaki et al. 2002a,b; Schaake-Koning et al. 1992; Curran et al. 2003). This combined treatment approach results in median survival times of 13–14 months, and in survival rates at 5 years as high as 15%–20%. These improvements have been achieved by using standard chemotherapeutic agents, primarily cisplatin-based drug combinations. Since direct comparison trials between induction and concurrent chemoradiotherapy have clearly demonstrated therapeutic superiority of the latter approach (Schaake-Koning et al. 1992; Curran et al. 2003), concurrent chemoradiotherapy can be regarded as the current standard of care

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for local-regionally advanced lung cancer. Still, the poor overall survival of lung cancer patients necessitates the introduction of treatment strategies that would further improve local tumor control, patient survival rate, and quality of life.

Many factors, known and unknown, limit therapeutic success of radiotherapy or chemoradiotherapy for lung cancer, with one major factor being the level of tolerance of normal tissues to the damage by these agents. Toxicities associated with chemotherapy and radiotherapy may limit the dose and duration of the treatment, adversely affect both short and long-term patient quality of life, be life-threatening, and increase costs of patient care. Normal tissue toxicities are more common and more serious after chemoradiotherapy than radiotherapy alone, and may be particularly ex- cessive in concurrent chemoradiotherapy. Because of the increased toxicity, the dose of chemotherapeutic agents in the setting of concurrent chemoradiotherapy is signiﬁ cantly reduced, which may lower drugs’ ability to exert their effects on both local-regional tumor and disseminated disease.

Because normal tissue toxicity is a major barrier to radiotherapy and chemoradiotherapy of lung cancer, ev- ery effort must be taken to avert or minimize the injury to critical normal tissues or other side effects of these treatments. Improvements are being sought primarily through better delivery of radiation therapy or the use of chemical or biological radio- or chemoprotective agents.

Technical improvements in radiotherapy include three- dimensional treatment planning, conformational radiotherapy, or the use of protons. These normal tissue spar- ing strategies may allow administration of higher doses of radiation, chemotherapeutic drugs, or both, directed towards achieving superior treatment outcome.

This chapter overviews a selection of relevant preclinical ﬁ ndings and limited clinical data on the use of radio- and chemo-protective agents to prevent or reduce injury to normal tissues that limit radiotherapy of lung cancer. We particularly focused our dis- cussion on protection with amifostine, and presented the results of our recent clinical trial. Additional in- formation can be found in other reviews on this topic (Murray and McBride 1996; Nieder et al. 2003).

2.2.6.2

Thiols as Radioprotective Agents

Both preclinical and clinical investigations on chemical protectors in radiotherapy have been dominated by thiols. The most effective compounds have those

with a sulfhydryl, –SH, group at one terminus and a strong basic function, an amino group, at the other terminus. Some of the important radioprotective thiols are listed in Table 2.2.6.1. The general structure of these aminothiols is H₂N(CH₂)_xNH(CH₂)_ySH, and among them, phosphorothioates (such as WR 2721, WR-3689, WR-151327) are the most effective and least toxic (Murray and McBride 1996). Various mechanisms have been proposed for the thiol-medi- ated radiation protection of normal tissues. Thiols (RSH) and their anions (RS-) rapidly bind to free radicals such as OH and prevent them from reacting with cellular DNA. This type of protection from DNA damage by scavenging free radicals is oxygen depen- dent (Travis 1984). Another mode of protection occurs via H-atom donation (the ﬁ xation-repair model).

Thiols compete with oxygen for radiation-induced DNA radicals. DNA radicals are “ﬁ xed” (not repaired) by reacting with oxygen and potentially harmful hy- droxyperoxides may be generated. However, DNA radicals can be chemically repaired when they react with thiols by donation of hydrogen (Durand 1983).

Furthermore, intracellular oxygen can be depleted as a result of thiol oxidation (Durand and Olive 1989) that would decrease the rate of oxygen-me- diated DNA damage ﬁ xation. Finally, thiols induce DNA packaging that may decrease accessibility of DNA sites to radiolytic attack. This mechanism may be oxygen independent and may explain the protection from densely ionizing radiation such as neutrons (Savoye et al. 1997).

2.2.6.2.1

Amifostine: Preclinical Findings

Amifostine (Ethyol) is a thiol-containing compound that has long been recognized for its strong radioprotective properties and has already been used in clinical trials (Brizel 2003). Amifostine does not readily cross the cell membrane because of its hydrophilicity. The drug is rapidly dephosphorylated to its active metabolite WR- 1065 and cleared from plasma with a half-life of 1–3 min following iv administration (Shaw et al. 1999a). In contrast to its brief systemic half-life, there is prolonged retention of the drug in normal tissues (Yuhas 1980). In the ﬁ rst 30 min following administration, drug uptake into normal tissues such as salivary gland, liver, kidney, heart, and bone marrow has been demonstrated to be up to 100-fold greater than in tumor tissues (Yuhas 1980). Bio-distribution studies show that the highest tissue levels of amifostine and its metabolites are found in salivary glands (Rasey et al. 1986).

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During the 1970s and 1980s extensive animal studies explored the ability of amifostine to protect a variety of normal tissues against acute and late radiation injury and whether the drug improves therapeutic ratio of radiotherapy. A radioprotective effect was observed for acute injury of the bone marrow, esophagus, jejunum, colon, hair follicles, testis, and immune system (Murray and McBride 1996; Milas et al.

1988). Amifostine was also a potent radioprotector of late responding tissues such as lung and subcutaneous tissues (Milas et al. 1988; Travis et al.

1985; Vujaskovic et al. 2002a,b; Lockhart 1990;

Hunter and Milas 1983). Protection of the lung was achieved against both single and fractionated radiation, and was assessed by biochemical testing such as reduction in hydroxyproline content of lung tissue and functional assays such as breathing fre- quency (Travis et al. 1985; Vujaskovic et al. 2002a).

Amifostine treatment was associated with reduction in accumulation of macrophages in irradiated lung and proﬁ brogenic cytokine activity (Vujaskovic et al. 2002b). Interestingly, while systemic application of amifostine was radioprotective for the lung tissue (Travis et al. 1985; Vujaskovic et al. 2002a,b), inhaled amifostine was ineffective (Lockhart 1990).

In contrast to the near universal protection of acutely responding tissues and lung, amifostine was not effective in protecting brain from radiation injury, which was attributed to the inability of the hydro- phyllic drug to cross the blood–brain barrier (Utley et al. 1984). Wide variation in the degree of radioprotection existed among various tissues, with protection factors for murine normal tissues ranging from 1.2 for hair follicles to greater than 2 for jejunum and

bone marrow (Murray and McBride 1996; Milas et al. 1988). The degree of radioprotection was de- pendent on the drug dose and time of administration in relation to radiation exposure. In general, higher doses of amifostine produced better protection up to a maximum dose of about 400 mg/kg (Murray and McBride 1996; Milas et al. 1982, 1988) Maximum radioprotection was achieved when amifostine was given 10–30 min before radiotherapy (Murray and McBride 1996; Milas et al. 1982, 1988). In addition to normal tissue radioprotection, a number of studies have examined whether amifostine protects tumors as well. Although some studies documented a small degree of tumor radioprotection, primarily of microscopic tumor foci, most studies showed no tumor protection (Murray and McBride 1996; Milas et al. 1982, 1988; Wasserman et al. 1981). Therefore, preclinical studies support the notion of selective or preferential normal tissue protection resulting in increased therapeutic gain of radiotherapy.

The mechanism of amifostine’s selective or preferential protection of normal tissues is related to several factors. Amifostine undergoes preferential rapid uptake into normal tissues but negligible or slower uptake into tumor tissues. While normal tissues actively concentrate amifostine against the concentration gradient, solid tumors generally absorb amifostine passively (Yuhas 1980). This selectivity results, in part, from differences in pH and alkaline phosphatase at the level of the capillary endothelium, both being higher in normal tissues compared to tumors (Yuhas 1980; Rasey et al. 1985, 1986). The acidic tumor microenvironment inhibits alkaline phosphatase necessary for uptake and conversion

Table 2.2.6.1. Radioprotective thiols and phosphorothioates. [Reprinted from Kirk-Othmer (1996), with permission]

CAS Register

Compound Number Structure

Thiols

Dithiothreitol (DTT) [27565-41-9] HSCH₂CH(OH)CH(OH)CH₂SH 2-Mercaptoethanol (WR-15504) [60-24-2] HOCH₂CH₂SH

Cysteamine (MEA, WR-347) [156-57-0] H₂NCH₂CH₂SH

2-((Aminopropyl)amino)ethanethiol [31098-42-7] H₂N(CH₃)₂NHCH₂CH₂SH (WR-1065)

WR-255591 [117062-90-5] CH₃NH(CH₂)₃NHCH₂CH₂SH

WR-151326 [120119-18-8] CH₃NH(CH₂)₃NH(CH₂)₃SH Phosphorothioates

WR-638 [3724-89-8] H₂NCH₂CH₂SPO₃H₂

WR-2721 [20537-88-6] H₂N(CH₂)₃NHCH₂CH₂SPO₃H₂ WR-3689 [20751-90-0] CH₃NH(CH₂)₃NHCH₂CH₂SPO₃H₂ WR-151327 [82147-31-7] CH₃NH(CH₂)₃NH(CH₂)₃SPO₃H₂

WR, Walter Reed Army Institute of Research.

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of amifostine to the active protective thiol, WR-1065 (Calabro-Jones et al. 1985), a condition absent in normal tissues. Once inside the cell, WR-1065 acts as a scavenger of oxygen free radicals (Ohnishi et al.

1992), which is reduced under hypoxic conditions commonly present in solid tumors. In addition, amifostine may be less available to tumors because of their defective vascular network.

Overall, a large body of preclinical data shows that amifostine preferentially protects the majority of normal tissues, including the lung, from the effects of DNA damaging agents, such as radiation. In addition to interaction with radiation, amifostine has been shown to exert independent antimetastatic and antiangiogenic activity (Grdina et al. 2002;

Giannopoulou and Papadimitriou 2003). Thus, these preclinical data provide a strong rationale for the clinical development of combined modality cancer treatment with amifostine and radiotherapy.

2.2.6.2.2

Amifostine: Clinical Studies

Clinical trials with amifostine began in the 1980s and showed that the drug is generally well tolerated. Its administration is associated with a number of transient side effects including nausea, vomiting, sneezing, mild somnolence, hypotension, a metallic taste during infusion, and occasional allergic reactions (Kligerman et al. 1988; Schuchter and Glick 1993). Hypotension appeared to be the most clinically signiﬁ cant side effect that could curtail treatment. A number of trials showed that amifostine reduces the severity of toxicity of radiotherapy or chemotherapy (Kligerman et al.

1988; Brizel et al. 2000; Kemp et al. 1996). Brizel et al. (2000) reported a randomized trial showing that amifostine reduces both severity and duration of xero- stomia in head and neck cancer patients treated with radiotherapy. This study led to FDA approval of amifostine for this clinical indication.

A number of clinical trials have been performed using amifostine in combination with chemoradiotherapy for lung cancer (Koukourakis et al. 2000;

Antonadou et al. 2001, 2003; Movsas et al. 2003;

Senzer 2002; Leong et al. 2003), including one at the University of Texas M. D. Anderson Cancer Center (MDACC) (Komaki et al. 2002a,b, 2004). Antonadou et al. (2001, 2003) conducted a randomized phase III trial of concurrent chemotherapy (either paclitaxel or carboplatin) and radiation treatment plus/minus daily amifostine given iv at 300 mg/m² 15–20 min before each fraction of radiotherapy and before chemother-

apy in patients with locally advanced lung cancer. The results showed that amifostine signiﬁ cantly reduced radiation-induced pneumonitis (>= grade 3 from 56.3% to 19.4%, p<0.002), and esophagitis (>= grade 3 from 84.4% to 38.9%, p<0.001) without compromising antitumor efﬁ cacy (Antonadou et al. 2003). Movsas et al. (2003) recently reported preliminary results of a phase III RTOG 98-01 trial in which 243 patients with stage II-IIIA/B NSCLC were treated with induction chemotherapy (paclitaxel and carboplatin) followed by concurrent chemotherapy and hyperfractionated radiotherapy (69.6 Gy with 1.2 Gy/fraction, BID).

Patients were randomized to receive amifostine i.v.

500 mg four times/week between the BID radiotherapy or no amifostine treatment. Although amifostine did not signiﬁ cantly reduce grade 3 or higher esophagitis, both weight loss from baseline and swallowing dysfunction were lower in the amifostine group.

At MDACC we investigated the ability of amifostine to reduce the severity and/or incidence of acute toxicities of concurrent chemotherapy and radiation therapy for NSCLC (Komaki et al. 2002a, 2004). A total of 64 patients with inoperable stage II or III NSCLC were treated with concurrent chemoradiotherapy. Both groups received thoracic radiation therapy (TRT) with 1.2 Gy/fraction, 2 fractions per day, 5 days per week for a total dose 69.6 Gy. All patients received oral etoposide (VP-16), 50 mg Bid, 30 min before TRT beginning day 1 for 10 days, repeated on day 29, and cisplatin 50 mg/m² iv on days 1, 8, 29, and 36. Patients in the study group received amifostine, 500 mg iv, twice weekly before chemoradiation (arm 1); patients in the control group received chemoradiation without amifostine (arm 2).

Patient and tumor characteristics were distributed equally in both groups. Of the 64 patients enrolled, 62 were evaluable (31 in arm 1, 31 in arm 2) with a mini- mum follow-up of 24 months. Important fi ndings from this study on the incidence and severity of a number of chemoradiotherapy-induced toxicities are shown in Figs. 2.2.6.1 and 2.2.6.2. As shown in Fig. 2.2.6.1, amifostine treatment increased the incidence of mild esophageal toxicity from 23% to 48%, but conversely it markedly reduced the incidence of severe esophageal toxicity from 35% to 16% (p=0.021). The reasons for this divergent effect of amifostine on mild and severe toxicity are not yet understood. Amifostine signifi - cantly reduced the incidence of constipation, pneumonitis, and neutropenic fever (Fig. 2.2.6.2). Of important note, severe, grade 3, pneumonitis occurred in 16% of patients treated with chemoradiotherapy alone but in no patients that received amifostine in addition to chemoradiotherapy. The most signifi cant side effect of amifostine was hypotension occurring in 65% of pa-

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tients, consistent with ﬁ ndings from other similar clinical studies. Figure 2.2.6.3 shows that amifostine had no signiﬁ cant effect on tumor response to chemoradiotherapy, as determined by percent of local regional control, percent of distant metastases free survival and overall patient survival. We concluded that amifostine reduced the severity and incidence of the acute esophageal, pulmonary, and hematologic toxicity resulting from concurrent cisplatin-based chemoradiotherapy, but had no apparent effect on tumor response to therapy. Another study from MDACC showed that amifostine can partially reverse the reduction of lung diffusion capacity caused by chemotherapy and/or radiotherapy (Gopal et al. 2002), further documenting amifostine-induced radioprotection of normal tissues during thoracic radiotherapy. The results of randomized clinical trials with iv amifostine in lung cancer are summarized in Table 2.2.6.2.

Although the iv route of amifostine administration has been most commonly used in clinical trials, the practical advantages of sc administration have led to a clinical trial directly comparing routs of administration. A study comparing the relative bioavailability of amifostine administered sc and iv was conducted in normal male volunteers. Amifostine was given either iv at a dose of 200 mg/m² or sc at a ﬁ xed dose of 500 mg. The sc dose resulted in an area under the concentration–time curve for the bound form of WR- 1065 of 68% compared to that after iv administration.

There was greater inter-patient variability in drug concentration following sc administration (Shaw et al. 1999b; Bonner and Shaw 2002).

Koukourakis et al. (2000) conducted a randomized Phase II study in 140 patients receiving radiotherapy to assess the feasibility, tolerance, and cyto- protective efﬁ cacy of sc amifostine. Patients (n=70)

received 500 mg of amifostine as a single sc injection 20 min prior to each radiotherapy fraction. The regimen was well tolerated, effectively reduced early toxicity of radiotherapy, and prevented treatment-induced delays. Patients reported a reduction in hypotension and nausea as compared with the iv administration. A phase III multi-center randomized trial to compare iv vs sc amifostine vs no amifostine in pa-

Fig. 2.2.6.1. Effect of amifostine on esophageal toxicity induced by chemoradiotherapy in patients with NSCLC. [Modiﬁ ed from Komaki et al. (2004) with permission]

Fig. 2.2.6.2. Effect of amifostine on constipation, pneumonitis, and neutropenic fever caused by chemoradiotherapy in patients with NSCLC. [Modiﬁ ed from Komaki et al. (2004) with permission]

Fig. 2.2.6.3. Kaplan-Meier survival curves showing the effect of amifostine on (a) overall survival rate, (b) locoregional (LR) tumor control, and (c) distant-metastasis (DM)-free survival after chemoradiotherapy in patients with NSCLC. [Modiﬁ ed from Komaki et al. (2004) with permission]

a

b

c

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tients with locally advanced NSCLC receiving concurrent chemoradiotherapy is ongoing. A phase II study of the efﬁ cacy of s.c. administration of amifostine in surgically resected NSCLC patients treated with post- operative radiotherapy is ongoing at MDACC.

2.2.6.3

Prostanoids, COX-2, and COX-2 Inhibitors

In response to physiological signals, stress or injury including radiation injury, cells produce prostanoids [prostaglandins (PGs) and thromboxanes (TBX)], a family of diverse, highly biologically active lipids de- rived from enzymatic metabolism of arachidonic acid by COX-1 or COX-2 enzymes. COX-1 is ubiquitous and responsible for prostanoid production in normal tissues where prostanoids exert numerous homeostatic physiological functions. In contrast, COX-2 is an in- ducible enzyme involved in prostaglandin production in pathologic states, particularly in inﬂ ammatory processes and cancer. COX-2 is induced by various factors including inﬂ ammatory cytokines (such as TNF-α, IL- 1β, and platelet activity factors), oncogenes, growth

factors, and hypoxia. Prostanoids play a role in the pathogenesis of various pathological states including infl ammation, where PGE2, a potent vasodilator and an immunosuppressive substance, is the major prostaglandin involved. PGE2, produced in abundance by pro-infl ammatory mononuclear cells such as macrophages, mediates the typical symptoms of infl ammation due to its vasodilatory action. This augments edema formation caused by substances that increase vascular permeability such as histamine. PGE2 is also involved in the development of erythema and heat at the site of infl ammation. Since radiation-induced lung injury is characterized by infl ammatory tissue reactions, PGE2 and other PGs, as well as pro-infl ammatory cytokines, are produced in injured tissue in abundance. Because different prostanoids have com- plementary or antagonistic activities, the fi nal biological effect on tissues depends on the balance of similar and opposing actions of the prostanoids involved.

Production of PGE2 and other pro-inﬂ ammatory prostanoids can be suppressed by non-steroidal anti- inﬂ ammatory drugs (NSAIDs), which inhibit both iso- forms of COX enzyme, or by selective COX-2 inhibitors.

Since selective COX-2 inhibitors do not inhibit prostanoid production in normal tissues, they are less toxic

Table 2.2.6.2. Randomized trials with amifostine in lung cancer. [From Komaki et al. (2004) with permission]

Reference Radiation dose Chemotherapy Amifostine dose Comments

Movsas 69.6 Gy @ 1.2 Gy Induction P+Cx2; 500 mg IV 4x/week No difference by NCI-CTC esophagitis et al. 2003 b.i.d. day 43 concurrent weekly C between b.i.d. Swallowing diaries

n=242 RT fractions (p=0.03) & weight loss

(p=0.05) favor amifostine

(Median survival, 15.6 and 15.8 months) Leong 60–66 Gy @ 2.0 Gy Induction P+Cx2; 740 mg/m2 with each Esophagitis grade 2–3:

et al. 2003 q.d. day 43 concurrent weekly P chemo (d 1, 22, 43, 50, 43% in amifostine, 70%

n=60 57, 64, 71, 78) in control (not signiﬁ cant)

(median survival, 12.5 and 14.5 months) Senzer 64.8 Gy @ 1.8 Gy Concurrent P+C q wk 500 mg IV before No difference in toxicity,

et al. 2002 q.d. day 1 x 7; gemcitabine & weekly chemo; 200 mg no survival data n=63 cisplatin x 3 after IV daily before RT (ongoing trial)

chemoradiation

Antonadou 55-60 Gy @ 2.0 Gy None 340 mg/m2 daily ↓ pneumonitis;

et al. 2001 q.d. before RT ↓ esophagitis

n=146 (no survival data)

Antonadou 55-60 Gy @ 2.0 Gy Concurrent weekly 300 mg/m2 daily ↓ esophagitis (p<0.001)

et al. 2003 q.d. P or C before Chemo/RT ↓ pneumonitis (p=0.009)

n=73 and RT (no survival data)

Komaki 69.6 Gy @1.2 Gy Concurrent cisplatin 500 mg IV 1st, 2nd ↓ degree of esophagitis, et al. 2004 b.i.d. day 1 IV d 1, 8, 29, 36 day each wk before ↓ pneumonitis n=62 Etoposide p.o. d 1-5 chemo & ↓ neutropenic fever

1st RT fraction 8–12, 29–33, 36–40

(median survival, 19 and 20 months)

P, paclitaxel; C, carboplatin; RT, radiation therapy; NCI-CTC, NCI common toxicity criteria.

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than commonly used NSAIDs. Interestingly, both prostanoids and their inhibitors have been reported to exert radioprotective actions on normal tissues. Exogenous administration of PGE2, other PGs, or PG analogs prior to irradiation of mice was shown to protect a variety of tissues including hematopoietic tissue, jejunal mucosa, dermis, and testis (reviewed in Hanson 1998; Milas and Hanson 1995). PGs vary widely in their radioprotective ability; however, the PG analog misoprostol was amongst the most effective. Paradoxically, inhibiting PGs by NSAIDs has also been shown to protect many tissues, including the lung, against radiation injury (Milas and Hanson 1995; Michalowski 1994). For example, Milas et al. (1992) reported that the NSAID indomethacin can protect mouse lung from radiation damage, but the protection was limited to the early pneumonitis phase of injury. Preliminary investigations in our laboratory using the selective COX-2 inhibitor SC-236 did not demonstrate signiﬁ cant protection from radiation-induced pneumonitis when the drug was administered a few days before and after lung irradiation. Subsequent experiments, using a different COX-2 inhibitor, celecoxib, provided suggestive evidence that giving the inhibitor during the development phase of acute pneumonitis may reduce either the latency or severity of lung injury. It should be emphasized that even in the absence of lung radioprotection by COX-2 inhibitors, therapeutic gain is still improved by their administration because of their potent enhancement of tumor radioresponse. The ability of COX-2 inhibitors to selec- tively enhance tumor radioresponse has been reviewed in detail elsewhere (Milas 2001; Milas et al. 2003b;

Choy and Milas 2003).

Corticosteroids are highly potent anti-inﬂ ammatory drugs used for symptomatic treatment of radiation-induced pneumonitis. They inhibit production of all prostanoids because, in addition to their ability to inhibit COX enzymes, they prevent release of arachidonic acid from membrane phospholipids by stimulating the generation and secretion of lipocor- tins. Ward et al. (1992a) showed that steroid administration to rats at the time of radiation delivery protected rats from lung interstitial edema, delayed or suppressed radiation-induced alveolitis, but did not affect development of pulmonary ﬁ brosis.

2.2.6.4

Growth Factors and Cytokines

Growth factors and cytokines play a critical role in pathogenesis of radiation injury, including that to

the lung. Radiation alters the magnitude and dy- namic activity of factors already present in affected tissues. Response to radiation occurs within minutes or hours after irradiation and can persist for days and months, inﬂ uencing the pathogenesis of both early and late radiation damage. The principal action of growth factors and cytokines is on cell and tissue proliferation, as well as cell loss. Hence, growth factors affect all of the major determinants of cell and tissue radioresponse: total number of clonogenic cells, cell cycle redistribution, cell repopulation, cellular repair mechanisms, and tissue microenvironment such as tumor hypoxia and acidity. Many growth factors may be affected upon tissue irradiation, those that have cytotoxic actions and those that have cytoprotective ability, so that the extent of tissue damage depends on the interaction of cytokines with similar or opposing activities. Involvement of growth factors and cytokines in pathogenesis of lung radiation damage is discussed in more detail in Chap. 11.6.

Since some growth factors and cytokines may act protectively, attempts have been made to protect tissues that are at risk from lung cancer radiotherapy.

Basic fi broblast growth factor (b-FGF) was found to protect endothelial cells both in vitro (Haimovitz- Friedman et al. 1991) and in vivo (Haimovitz- Friedman et al. 1991; Fuks et al. 1994) from radiation. To confer radiation resistance in vitro, b-FGF had to be present at the time of radiation exposure and/or within several hours after irradiation. This protective effect was abolished by treatment with anti-b-FGF an- tibodies. The radioprotective effect of b-FGF was attributed to its ability to increase cellular repair. A subsequent study by the same group (Fuks et al. 1994) showed that mice could be protected from lethal doses of whole lung irradiation if given iv b-FGF immedi- ately before or within 2 h after irradiation. The effect was attributed to the protection of endothelial cells against radiation-induced apoptosis. Histology of irradiated lung tissue, but not of lungs exposed to both b-FGF and radiation, showed apoptotic changes in the endothelial cell lining of the pulmonary microvascu- lature within 6–8 h after radiation exposure. Also, his- tological features of radiation-induced pneumonitis were absent in mice treated with b-FGF. These results were not confi rmed in a subsequent study by Tee and Travis (1995) that assessed the radioprotective action of b-FGF in two strains of mice having different susceptibilities to radiation-induced lung injury. The reasons for the discrepancy are unclear, but some differences in experimental conditions such as radiation dose, fi eld size of radiation, and mouse strain could have accounted for this disparity.

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Keratinocyte growth factor (KGF) is another cell growth factor that has been investigated for its ability to protect against radiation-induced lung damage.

Although a member of the FGF family (FGF-7), KGF’s cell growth stimulatory activity is conﬁ ned to epithelial cells (Rubin et al. 1989; Miki et al. 1992). KGF was shown to be a good stimulator of proliferation of type II pneumocytes in vitro and in vivo (Panos et al. 1993;

Ulich et al. 1994), a type of cells considered to play an important role in repair of injured lung tissue. Yi et al.

(1996) showed that intratracheal administration of KGF to rats 2 or 3 days before exposure of rats to 18 Gy bilat- eral thoracic irradiation reduced severity of radiation- induced pneumonitis and fi brosis observed histologically. However, there was no signifi cant improvement in rat survival. In contrast, KGF was highly effective both in preventing development of bleomycin-induced fi brosis and in improving the survival of treated animals. Signifi cant protection was also rendered against the damage infl icted by the combined bleomycin plus radiation treatment. A more recent study (Terry et al.

2004) showed that a single intratracheal administration of rHuKGF to normal mice increased proliferation of alveolar epithelial cells 3–7 days later. This treatment afforded signiﬁ cant protection against lethality from radiation-induced pneumonitis when the mice were irradiated at day 7 after administration of rHuKGF.

Regarding radioprotective abilities of cytokines, it is worth mentioning that sc administration of recombinant IL-11 (rIL-11) rendered signiﬁ cant protection to mice from fatal thoracic irradiation (Redlich et al. 1996). The observed radioprotection was attributed to the rIL-11-induced inhibition of radiation- induced expression of TNF mRNA as well as TNF production by macrophages.

2.2.6.5 Pentoxifylline

Pentoxifylline (Trental), a methylxanthine derivative, is hemorheologic agent capable of reducing or ameliorat- ing late radiation sequelae. In humans, pentoxifylline is used to treat persistent soft tissue ulcerations and necrosis. It has a variety of physiological activities including inhibition of platelet aggregation, regulation of tissue damaging cytokines such as tumor necrosis factor alpha (TNF_), and enhances blood ﬂ ow in injured mi- crovasculature. The drug may increase radioresponse of solid tumors by increasing tumor oxygenation (Lee et al. 2000), and as such was tested in a clinical phase III trial in NSCLC in combination with radiotherapy

(Kwon et al. 2000). This study showed that pentoxifylline was modestly effective, increasing median time to relapse by 2 months and median survival time from 7 to 18 months. Though pentoxifylline has been shown to modestly improve tumor radioresponse, it has most often been used to reduce normal tissue radiation injury. Preclinical studies in experimental animals have, in general, shown pentoxifylline to be radioprotective but the degree of protection was highly variable. As an illustration, Lefaix et al. (1999) reported striking regression of subcutaneous ﬁ brosis induced by radiation to the skin surface of pigs using a combination of pentoxifylline and alpha-tocopherol (vitamin E). On the other hand, pentoxifylline has also been shown to have little or no effect on acute skin or lung injuries (Dion et al. 1989; Koh et al. 1995; Rube et al. 2002;

Ward et al. 1992b). With respect to lung injury, pentoxifylline inhibited the radiation-induced increase in TNF_ mRNA during the acute phase of radiation injury, pneumonitis, but the impact of this biochemical change on lung injury was unclear (Rube et al. 2002). In another study (Ward et al. 1992b), pentoxifylline was found to further increase radiation-induced production of prostanoids (PGI2 and TXA2), while decreasing endothelial dysfunction accompanied by increases in lung wet weight, protein, and hydroxyproline content in the irradiated lung. A recent randomized clinical trial, however, using prophylactic pentoxifylline showed a signiﬁ cant reduction in both early and late radiation- induced lung toxicities in patients with breast and lung cancer (Ozturk et al. 2004).

2.2.6.6

Angiotensin Converting Enzyme (ACE) Inhibitors

Angiotensin converting enzyme (ACE) converts angiotensin I to angiotensin II, which is a potent va- soconstrictor and hypertensive factor. Captopril is an inhibitor of ACE that has been shown to protect against radiation injury of a number of tissues including the lung. In addition to inhibition of ACE, captopril is a free radical scavenger (Chopra et al. 1989) and exhibits superoxide dismutase (SOD)-like activity (Roberts and Robinson 1995). Captopril reduces radiation-induced pulmonary endothelial dysfunction (Ward et al. 1988), pulmonary ﬁ brosis (Ward et al.

1990a, 1992c), and delays radiation-induced pulmonary arterial hypoperfusion in rats (Graham et al.

1988). Moreover, ACE inhibitor prophylaxis in rats receiving whole lung radiation was found to reduce radiation-induced activation of ACE, plasminogen ac-

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tivator, and production of prostaglandins, and throm- boxane (Ward et al. 1988). When added to the feed after irradiation, captopril reduced early lung reaction in rats receiving fractionated hemithoracic irradiation (Ward et al. 1993). In addition to pulmonary protection, ACE inhibition also protects against radiation injury of other tissues including kidney (Moulder et al. 1993), skin (Ward et al. 1990b), jejunum (Yoon et al. 1994), and heart (Yarom et al. 1993). With respect to the heart, Yarom et al. (1993) showed that captopril ameliorated the decrease in capillary function, increase in mast cells, ﬁ brosis, number of atrial granules, and changes in nerve terminals, but it failed to prevent the progressive functional deterioration of the heart following irradiation. Mechanisms of captopril-medi- ated radioprotection are not fully understood, but are at least partially related to its antihypertensive activity and its thiol-like function.

Based on promising preclinical observations on radioprotection by ACE inhibitors, several clinical studies were performed. Wang et al. (2000) reported a retrospective clinical study of ACE inhibitors given for the management of hypertension in patients with lung cancer treated with deﬁ nitive radiotherapy. The study showed ACE inhibitors given at a dose within the range used to treat hypertension did not decrease the incidence or delay the onset of symptomatic radiation pneumonitis. Currently, a phase II RTOG randomized clinical trial using captopril is ongoing. The primary goal of this study is to test whether captopril given after completion of radiotherapy can reduce the incidence or severity of pulmonary damage after aggressive deﬁ nitive chemoradiotherapy.

2.2.6.7

Radioprotective Gene Therapy:

Superoxide Dismutase (SOD)

The manganese superoxide dismutase (MnSOD) lo- cated within the mitochondria is one of nature’s most effi cient catalysts. The enzyme protects redox ma- chinery within the mitochondria from the superoxide radical produced during normal respiration. In many pathological conditions, such as infl ammation caused by radiation-induced free radical damage, superoxide is abundantly produced and may overwhelm the cell’s ability to effi ciently remove thus leading to tissue injury.

Via its antioxidant activity, MnSOD inactivates superoxide and hence has potential to protect against free-radical induced injury. Early studies showed that systemic administration of SOD can prevent radiation injury

(Petkau 1987) and can even reduce preexisting radiation-induced ﬁ brosis (Delanian et al. 1994). When given prior to radiation the activity of SOD has generally been attributed to its radical scavenging effects, whereas when given after radiation the effects are most likely related to its anti-inﬂ ammatory and or immuno- stimulatory properties (Murray and McBride 1996).

Another SOD, recombinant CuZnSOD was shown to protect the lung of hamsters from radiation-induced damage as evidenced by the absence of severe histo- pathologic tissue changes 4–16 weeks after irradiation and the prevention of elevation of total protein content in bronchoalveolar lavage (Breuer et al. 2000).

More recently, a novel approach has been advanced for radioprotective gene therapy using the antioxidant manganese superoxide dismutase delivered to specifi c target organs such as lung and esophagus by gene transfer vectors including plasmid/liposomes (PL) and adenovirus (Greenberger et al. 2003). Radiation protection by MnSOD transgene overexpression at the cellular level has been demonstrated to be localized to the mitochondrial membrane. Intraesophageal administration of MnSOD-PL prior to irradiation induces transgene expression for 48–72 h, and an associated decrease in radiation-induced expression of infl ammatory cytokine mRNA and protein and esophagitis (Epperly et al. 2001, 2003). Intratracheal injection of adenovirus containing MnSOD protected against radiation-induced organizing alveolitis in mice (Epperly et al. 1999). In addition, intratracheal MnSOD-PL gene therapy reduced radiation induced infl ammatory cytokines without rendering protection to orthotopic Lewis lung cancer (Guo et al. 2003). Preclinical animal studies suggested that radioprotective gene therapy reduces the radiation toxicities and may facilitate dose escalation protocols to improve the therapeutic ratio of lung cancer radiotherapy.

However, the efﬁ cacy and speciﬁ city of this approach need further investigation. Application of MnSOD-PL gene therapy in the setting of fractionated chemo-radiotherapy is being tested in clinical trials for prevention of esophagitis in patients with non-small cell lung cancer.

The gene therapy approach to speciﬁ cally deliver agents to targeted tissues is not limited to MnSOD but has high potential for delivery of a wide array of agents including both cytotoxic and radioprotective agents.

2.2.6.8

Concluding Remarks

Normal tissue damage remains a major limiting factor in cancer radiotherapy, and chemoradiotherapy

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where the improvement in tumor control and survival of patients is accompanied by increased rate and severity of treatment related toxicity. For many years scientists have explored various approaches to minimize damage to tissues, including the use of chemical and biological radioprotective agents. As elaborated in this chapter, many of these agents exert signiﬁ cant radioprotection and chemoprotection in experimental animal models and some of them have been tested in clinical trials. Amifostine has undergone the most extensive investigation, both preclinical and clinical.

A number of clinical trials, including a recent one from MDACC described in Sect. 2.2.6.2.2, provided encouraging results both with respect to reduction of the incidence and/or severity of esophagitis and pneumonitis. Agents discussed have complex mechanisms of action, and affect a variety of radiation-induced tissue reactions both directly and indirectly.

Radiation elicits the release of many substances, such as growth factors, cytokines and prostanoids, which can have both radioprotective and radioenhancing properties. Since the ﬁ nal outcome of treatment criti- cally depends on the balance between these compet- ing processes, the use of radioprotective agents may act on only some of the many factors involved. This is likely one of the reasons for the inconsistency in the literature on radioprotection. Rapid achievements in recombinant technology and genetic engineering are opening possibilities to upregulate or downregulate cellular expressions of diverse factors involved in tissue responses to radiation and to select appro- priate factors to achieve a predetermined response.

For example, redirecting the actions or optimizing the concentrations of a given response factor may become useful in increasing the therapeutic ratio of radiotherapy by enhancing tumor radioresponse, or by reducing damage of normal tissues with radioprotectors.

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